Antigen binding proteins that cross-neutralize staphylococcus aureus toxins and uses thereof

Abstract

The present application relates to an isolated antigen binding protein targeting Staphylococcus aureus and proteins derived thereof, which are capable of having cross-neutralizing activity against multiple toxins of Staphylococcus aureus. The present application also discloses the use of the above-mentioned isolated antigen binding protein and proteins derived thereof in the field of treating and / or preventing Staphylococcus aureus infection.

Description

Technical Field

[0001] This application relates to the field of biomedicine, specifically to an antigen-binding protein that specifically binds to Staphylococcus aureus toxin. Background Technology

[0002] Staphylococcus aureus belongs to the genus Staphylococcus and is an important Gram-positive pathogenic bacterium that can cause purulent infections, pneumonia, pseudomembranous colitis, pericarditis, and other local infections, as well as systemic infections such as sepsis and septicemia. Staphylococcus aureus primarily causes disease by producing toxins and invasive enzymes, including hemolysins, leukocidins, enterotoxins, and coagulases.

[0003] Hemolysins can be classified into four types based on their antigens: α-hemolysins, β-hemolysins, γ-hemolysins, and δ-hemolysins. γ-hemolysins are universally expressed in all Staphylococcus aureus strains, and their gene products can produce two types of toxins: HlgAB and HlgCB. γ-hemolysins are highly effective at killing human immune cells, including neutrophils, lymphocytes, and macrophages, and can also destroy human erythrocytes. Leukocidins (PVL) are extracellular toxins produced by Staphylococcus aureus, belonging to the membrane-burr-toxin family, and consist of two subunits, "S" and "F". The F and S components of Staphylococcus aureus toxins can form both homologous and non-homologous pairs. Due to the abundance and heterogeneity of toxin families, inactivation of a single component is unlikely to be effective against Staphylococcus aureus infection. Therefore, there is an urgent need to develop drugs that can effectively combat different two-component Staphylococcus aureus strains. Summary of the Invention

[0004] This application provides an isolated antigen-binding protein having one or more of the following properties: 1) specifically binding to Staphylococcus aureus toxins; 2) specifically binding to one or more of the following Staphylococcus aureus toxins: HClA, HClB, LukD, and LukF; 3) neutralizing the activity of one or more Staphylococcus aureus toxins; and 4) treating diseases, symptoms, and / or complications caused by Staphylococcus aureus. The isolated antigen-binding protein described in this application exhibits in vitro affinity and neutralizing activity against both groups of toxins, as well as in vivo pharmacodynamic activity in animals, and provides protection against Staphylococcus aureus-induced infection in mice.

[0005] On one hand, this application provides an isolated antigen-binding protein comprising at least one CDR in the variable region VH of the antibody heavy chain, said VH comprising the amino acid sequence shown in SEQ ID NO:53.

[0006] In some embodiments, the isolated antigen-binding protein comprises HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:1.

[0007] In some embodiments, the isolated antigen-binding protein comprises HCDR2, which comprises the amino acid sequence shown in SEQ ID NO:2.

[0008] In some embodiments, the isolated antigen-binding protein comprises HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:3.

[0009] In some embodiments, the isolated antigen-binding protein comprises HCDR1, HCDR2, and HCDR3, wherein HCDR1 comprises the amino acid sequence shown in SEQ ID NO:3, HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2, and HCDR3 comprises the amino acid sequence shown in SEQ ID NO:1.

[0010] In some embodiments, the isolated antigen-binding protein comprises H-FR1, the C-terminus of which is directly or indirectly linked to the N-terminus of HCDR1, and the H-FR1 comprises the amino acid sequence shown in SEQ ID NO:45.

[0011] In some embodiments, the isolated antigen-binding protein H-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:10, SEQ ID NO:14, and SEQ ID NO:18.

[0012] In some embodiments, the isolated antigen-binding protein comprises H-FR2, which is located between HCDR1 and HCDR2, and which comprises the amino acid sequence shown in SEQ ID NO:46.

[0013] In some embodiments, the isolated antigen-binding protein H-FR2 comprises the amino acid sequence shown in any one of SEQ ID NO:11, SEQ ID NO:15, and SEQ ID NO:19.

[0014] In some embodiments, the isolated antigen-binding protein comprises H-FR3, which is located between HCDR2 and HCDR3, and the H-FR3 comprises the amino acid sequence shown in SEQ ID NO:47.

[0015] In some embodiments, the isolated antigen-binding protein H-FR3 comprises the amino acid sequence shown in any one of SEQ ID NO:12, SEQ ID NO:16, and SEQ ID NO:20.

[0016] In some embodiments, the isolated antigen-binding protein comprises H-FR4, the N-terminus of which is linked to the C-terminus of HCDR3, and the H-FR4 comprises the amino acid sequence shown in SEQ ID NO:48.

[0017] In some embodiments, the isolated antigen-binding protein H-FR4 comprises the amino acid sequence shown in SEQ ID NO:13 or SEQ ID NO:17.

[0018] In some embodiments, the isolated antigen-binding protein comprises H-FR1, H-FR2, H-FR3, and H-FR4, wherein H-FR1 comprises the amino acid sequence shown in SEQ ID NO:45, H-FR2 comprises the amino acid sequence shown in SEQ ID NO:46, H-FR3 comprises the amino acid sequence shown in SEQ ID NO:47, and H-FR4 comprises the amino acid sequence shown in SEQ ID NO:48.

[0019] In some embodiments, the isolated antigen-binding protein comprises H-FR1, H-FR2, H-FR3, and H-FR4, wherein H-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:10, SEQ ID NO:14, and SEQ ID NO:18, H-FR2 comprises the amino acid sequence shown in any one of SEQ ID NO:11, SEQ ID NO:15, and SEQ ID NO:19, H-FR3 comprises the amino acid sequence shown in any one of SEQ ID NO:12, SEQ ID NO:16, and SEQ ID NO:20, and H-FR4 comprises the amino acid sequence shown in SEQ ID NO:13 or SEQ ID NO:17.

[0020] In some embodiments, the isolated antigen-binding protein comprises H-FR1, H-FR2, H-FR3, and H-FR4 selected from any of the following groups:

[0021] 1) H-FR1 contains the amino acid sequence shown in SEQ ID NO:10, H-FR2 contains the amino acid sequence shown in SEQ ID NO:11, H-FR3 contains the amino acid sequence shown in SEQ ID NO:12, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:13;

[0022] 2) H-FR1 contains the amino acid sequence shown in SEQ ID NO:14, H-FR2 contains the amino acid sequence shown in SEQ ID NO:15, H-FR3 contains the amino acid sequence shown in SEQ ID NO:16, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17;

[0023] 3) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:15, H-FR3 contains the amino acid sequence shown in SEQ ID NO:16, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17;

[0024] 4) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:19, H-FR3 contains the amino acid sequence shown in SEQ ID NO:16, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17;

[0025] 5) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:15, H-FR3 contains the amino acid sequence shown in SEQ ID NO:20, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17; and

[0026] 6) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:19, H-FR3 contains the amino acid sequence shown in SEQ ID NO:20, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17.

[0027] In some embodiments, the isolated antigen-binding protein comprises VH, which contains the amino acid sequence shown in SEQ ID NO:53.

[0028] In some embodiments, the VH of the isolated antigen-binding protein comprises the amino acid sequence shown in any one of SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0029] In some embodiments, the isolated antigen-binding protein includes the constant region of the antibody heavy chain.

[0030] In some implementations, the heavy chain constant region is derived from the human IgG constant region.

[0031] In some implementations, the heavy chain constant region is derived from the human IgG1 heavy chain constant region.

[0032] In some embodiments, the heavy chain constant region comprises the amino acid sequence shown in SEQ ID NO:42.

[0033] In some embodiments, the isolated antigen-binding protein comprises an antibody heavy chain containing the amino acid sequence shown in any one of SEQ ID NO:55, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60, and SEQ ID NO:61.

[0034] In some embodiments, the isolated antigen-binding protein comprises at least one CDR in the variable region (VL) of the antibody light chain, the VL comprising the amino acid sequence shown in SEQ ID NO:54.

[0035] In some embodiments, the isolated antigen-binding protein comprises LCDR3, which contains the amino acid sequence shown in SEQ ID NO:4.

[0036] In some embodiments, the isolated antigen-binding protein comprises LCDR2, which contains the amino acid sequence shown in SEQ ID NO:5.

[0037] In some embodiments, the isolated antigen-binding protein comprises LCDR1, which contains the amino acid sequence shown in SEQ ID NO:44.

[0038] In some embodiments, the LCDR1 comprises the amino acid sequence shown in any one of SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, and SEQ ID NO:9.

[0039] In some embodiments, the isolated antigen-binding protein comprises LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises the amino acid sequence shown in SEQ ID NO:44, LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5, and LCDR3 comprises the amino acid sequence shown in SEQ ID NO:4.

[0040] In some embodiments, the isolated antigen-binding protein comprises LCDR1, LCDR2, and LCDR3, wherein LCDR1 comprises the amino acid sequence shown in any one of SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, and SEQ ID NO:9, LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5, and LCDR3 comprises the amino acid sequence shown in SEQ ID NO:4.

[0041] In some embodiments, the isolated antigen-binding protein comprises L-FR1, the C-terminus of which is directly or indirectly linked to the N-terminus of LCDR1, and the L-FR1 comprises the amino acid sequence shown in SEQ ID NO:49.

[0042] In some embodiments, the isolated antigen-binding protein L-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:21, SEQ ID NO:25, and SEQ ID NO:29.

[0043] In some embodiments, the isolated antigen-binding protein comprises L-FR2, which is located between LCDR1 and LCDR2, and the L-FR2 comprises the amino acid sequence shown in SEQ ID NO:50.

[0044] In some embodiments, the isolated antigen-binding protein L-FR2 comprises the amino acid sequence shown in SEQ ID NO:22 or SEQ ID NO:26.

[0045] In some embodiments, the isolated antigen-binding protein comprises L-FR3 located between LCDR2 and LCDR3, and the L-FR3 comprises the amino acid sequence shown in SEQ ID NO:51.

[0046] In some embodiments, the isolated antigen-binding protein L-FR3 comprises the amino acid sequence shown in SEQ ID NO:23 or SEQ ID NO:27.

[0047] In some embodiments, the isolated antigen-binding protein comprises L-FR4, the N-terminus of which is directly or indirectly linked to the C-terminus of LCDR3, and the L-FR4 comprises the amino acid sequence shown in SEQ ID NO:52.

[0048] In some embodiments, the isolated antigen-binding protein L-FR4 comprises the amino acid sequence shown in SEQ ID NO:24 or SEQ ID NO:28.

[0049] In some embodiments, the isolated antigen-binding protein comprises L-FR1, L-FR2, L-FR3, and L-FR4, wherein L-FR1 comprises the amino acid sequence shown in SEQ ID NO:49, L-FR2 comprises the amino acid sequence shown in SEQ ID NO:50, L-FR3 comprises the amino acid sequence shown in SEQ ID NO:51, and L-FR4 comprises the amino acid sequence shown in SEQ ID NO:52.

[0050] In some embodiments, the isolated antigen-binding protein comprises L-FR1, L-FR2, L-FR3, and L-FR4, wherein L-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:21, SEQ ID NO:25, and SEQ ID NO:29, L-FR2 comprises the amino acid sequence shown in SEQ ID NO:22 or SEQ ID NO:26, L-FR3 comprises the amino acid sequence shown in SEQ ID NO:23 or SEQ ID NO:27, and L-FR4 comprises the amino acid sequence shown in SEQ ID NO:24 or SEQ ID NO:28.

[0051] In some embodiments, the isolated antigen-binding protein comprises L-FR1, L-FR2, L-FR3, and L-FR4 selected from any of the following groups:

[0052] 1) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:21, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:22, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:23, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:24;

[0053] 2) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:25, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:26, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:27, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:28; and

[0054] 3) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:29, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:26, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:27, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:28.

[0055] In some embodiments, the isolated antigen-binding protein comprises a VL containing the amino acid sequence shown in SEQ ID NO:54.

[0056] In some embodiments, the VL comprises the amino acid sequence shown in any one of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39 and SEQ ID NO:40.

[0057] In some embodiments, the isolated antigen-binding protein comprises a constant region of the antibody light chain.

[0058] In some implementations, the light chain constant region is derived from the human Igκ constant region.

[0059] In some embodiments, the light chain constant region comprises the amino acid sequence shown in SEQ ID NO:43.

[0060] In some embodiments, the isolated antigen-binding protein comprises an antibody light chain, the light chain comprising the amino acid sequence shown in any one of SEQ ID NO:56, SEQ ID NO:62, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:65 and SEQ ID NO:66.

[0061] In some embodiments, the isolated antigen-binding protein comprises any one of the following groups of VH and VL:

[0062] 1) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0063] 2) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0064] 3) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0065] 4) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0066] 5) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains {SEQ ID NO:40}

[0067] The amino acid sequence shown;

[0068] 6) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0069] 7) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0070] 8) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0071] 9) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0072] 10) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0073] 11) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0074] 12) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0075] 13) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0076] 14) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0077] 15) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0078] 16) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0079] 17) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0080] 18) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0081] 19) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0082] 20) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0083] 21) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0084] 22) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0085] 23) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0086] 24) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0087] 25) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0088] 26) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0089] 27) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0090] 28) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0091] 29) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0092] 30) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0093] 31) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0094] 32) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0095] 33) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0096] 34) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0097] 35) The VH comprises the amino acid sequence shown in SEQ ID NO:35, and the VL comprises the amino acid sequence shown in SEQ ID NO:40; and

[0098] 36) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:41.

[0099] In some embodiments, the isolated antigen-binding protein comprises an antibody or an antigen-binding fragment thereof.

[0100] In some embodiments, the antigen-binding fragment includes Fab, Fab', Fv fragments, F(ab')2, F(ab)2, scFv, di-scFv, and / or dAb.

[0101] In some embodiments, the antibody is selected from one or more of the group consisting of monoclonal antibodies, chimeric antibodies, humanized antibodies, and fully human antibodies.

[0102] In some embodiments, the isolated antigen-binding protein is capable of specifically binding to Staphylococcus aureus toxin.

[0103] In some embodiments, the Staphylococcus aureus toxin comprises leukocidin.

[0104] In some embodiments, the Staphylococcus aureus toxin is selected from one or more of the group consisting of HlgA, HlgB, LukD, and LukF.

[0105] In some embodiments, the isolated antigen-binding protein is capable of neutralizing the activity of one or more Staphylococcus aureus virulence factors selected from the group consisting of HlgA, HlgB, LukD, and LukF.

[0106] In some embodiments, the isolated antigen-binding protein is capable of treating diseases and / or conditions caused by Staphylococcus aureus.

[0107] In some embodiments, the disease and / or condition includes complications of diseases and / or conditions caused by Staphylococcus aureus.

[0108] On the other hand, this application also provides a polypeptide molecule comprising the isolated antigen-binding protein.

[0109] In some embodiments, the polypeptide molecule comprises a fusion protein.

[0110] On the other hand, this application also provides an immunoconjugate comprising the isolated antigen-binding protein.

[0111] On the other hand, this application also provides a nucleic acid molecule that encodes the isolated antigen-binding protein or the polypeptide molecule.

[0112] On the other hand, this application also provides a vector containing the nucleic acid molecule.

[0113] On the other hand, this application also provides cells containing the nucleic acid molecules or the vector.

[0114] On the other hand, this application also provides pharmaceutical compositions comprising the isolated antigen-binding protein, the polypeptide molecule, the immunoconjugate, the nucleic acid molecule, the carrier and / or the cell, and optionally a pharmaceutically acceptable carrier.

[0115] On the other hand, this application also provides a method for preparing the isolated antigen-binding protein, the method comprising culturing the cells under conditions that cause the antigen-binding protein to be expressed.

[0116] On the other hand, this application also provides the use of the isolated antigen-binding protein, the polypeptide molecule, the immunoconjugate, the nucleic acid molecule, the carrier, the cell and / or the pharmaceutical composition in the preparation of a medicament for the prevention and / or treatment of diseases and / or conditions.

[0117] In some embodiments, the disease and / or symptoms and their complications are caused or mediated by Staphylococcus aureus.

[0118] In some implementations, the disease and / or condition includes complications of the disease and / or condition caused or mediated by Staphylococcus aureus.

[0119] In some implementations, the disease and / or symptom includes sepsis and / or bacteremia.

[0120] On the other hand, the isolated antigen-binding protein, the polypeptide molecule, the immunoconjugate, the nucleic acid molecule, the carrier, the cell, and / or the pharmaceutical composition described in this application may be used alone or in combination with other drugs.

[0121] On the other hand, this application also provides a method for detecting Staphylococcus aureus leukocidin in a sample, the method comprising administering the isolated antigen-binding protein, the polypeptide molecule, the immunoconjugate, the nucleic acid molecule, the carrier, the cell and / or the pharmaceutical composition.

[0122] On the other hand, this application also provides a kit for detecting Staphylococcus aureus leukocidin in a sample, the kit comprising the administration of the isolated antigen-binding protein, the polypeptide molecule, the immunoconjugate, the nucleic acid molecule, the carrier, the cell and / or the pharmaceutical composition.

[0123] On the other hand, this application also provides the use of the isolated antigen-binding protein, the polypeptide molecule, the immunoconjugate, the nucleic acid molecule, the carrier, the cell and / or the pharmaceutical composition in the preparation of a kit for detecting the presence and / or content of Staphylococcus aureus leukocidin in a sample.

[0124] Other aspects and advantages of this application will readily be apparent to those skilled in the art from the detailed description below. Only exemplary embodiments of this application are shown and described in the following detailed description. As will be appreciated by those skilled in the art, the content of this application enables them to make modifications to the disclosed specific embodiments without departing from the spirit and scope of the invention to which this application pertains. Accordingly, the descriptions in the accompanying drawings and specification of this application are merely exemplary and not restrictive. Attached Figure Description

[0125] The specific features of the invention involved in this application are shown in the appended claims. The features and advantages of the invention can be better understood by referring to the exemplary embodiments and drawings described in detail below. A brief description of the drawings is as follows:

[0126] Figure 1 The AD display shows the binding activity of the antigen-binding protein described in this application with Staphylococcus aureus leukocidin.

[0127] Figure 2 The AC display shows the in vitro neutralizing activity of the antigen-binding protein described in this application against Staphylococcus aureus leukocidins.

[0128] Figure 3 This application demonstrates the protective effect of the antigen-binding protein described herein against Staphylococcus aureus infection in mice.

[0129] Figure 4 The results shown are the affinity test results between the antigen-binding protein described in this application and the Staphylococcus aureus leukocidin HlgB.

[0130] Figure 5 The results shown are the affinity test results between the antigen-binding protein described in this application and the Staphylococcus aureus leukocidin HlgB.

[0131] Figure 6 The results shown are the affinity test results between the antigen-binding protein described in this application and the Staphylococcus aureus leukocidin HlgB.

[0132] Figure 7 The AD display shows the affinity test between the antigen-binding protein described in this application and Staphylococcus aureus leukocidin.

[0133] Figure 8 The results shown are from a mouse sepsis model induced by Staphylococcus aureus α-toxin, used to detect the therapeutic effect of the antigen-binding protein described in this application.

[0134] Figure 9 The results shown are from an evaluation of the anti-infective pharmacodynamics of the antigen-binding protein described in this application using a mouse bacteremia model induced by Staphylococcus aureus. Detailed Implementation

[0135] The following specific embodiments illustrate the implementation of the invention. Those skilled in the art can easily understand other advantages and effects of the invention from the content disclosed in this specification.

[0136] Terminology Definition

[0137] In this application, the term "Staphylococcus aureus" is used interchangeably with "Staphylococcus aureus," "S. aureus," "Staphylococcus pyogenes," and "S. aureus," which generally belong to Gram-positive bacteria. In this application, the term can encompass any type and form of Staphylococcus aureus and its variants, homologs, and functionally active fragments. The term "Staphylococcus aureus" can also include any toxins secreted by Staphylococcus aureus and its variants, homologs, and functionally active fragments. The term "Staphylococcal toxin" is used interchangeably with "Staphylococcal virulence factor." The term "leukocidin" includes, but is not limited to, S-components such as LukS-PV, HlgA (γ-hemolysin), HlgC (γ-hemolysin), LukE from Staphylococcus aureus, and LukS-I (from Staphylococcus intermedius); and F-components such as LukF-PV, LukF'-PV, HlgB (γ-hemolysin), LukD from Staphylococcus aureus, and LukF-I (from Staphylococcus intermedius). This application may cover one or more of various leukocidins, as well as variants, homologs, analogs, and functionally active fragments of leukocidins.

[0138] In this application, the term "isolated" generally refers to something obtained artificially from its natural state. If a substance or component is found in nature as an "isolated" substance, it may be due to an alteration of its natural environment, the isolation of the substance from its natural environment, or both. For example, a certain unisolated polynucleotide or polypeptide may naturally exist in the body of a living animal, and a high-purity identical polynucleotide or polypeptide isolated from this natural state is called isolated. The term "isolated" does not exclude the presence of artificial or synthetic substances, nor does it exclude the presence of other impurities that do not affect the activity of the substance.

[0139] In this application, the term "isolated antigen-binding protein" generally refers to a protein that has antigen-binding ability after being removed from its native state. This "isolated antigen-binding protein" may include a portion that binds to an antigen and, optionally, allow the antigen-binding portion to employ a framework or scaffold portion that promotes the antigen-binding portion's conformation for binding to the antigen. The antigen-binding protein may include, for example, an antibody-derived protein framework region (FR) or an alternative protein framework region or artificial framework region having a transplanted CDR or CDR derivative. Such frameworks include, but are not limited to, antibody-derived framework regions containing mutations introduced, for example, to stabilize the three-dimensional structure of the antigen-binding protein, and fully synthetic framework regions containing, for example, biocompatible polymers. See, for example, Korndorfer et al., 2003, Proteins: Structure, Function, and Bioinformatics, 53(1):121-129 (2003); Roque et al., Biotechnol. Prog. 20:639-654 (2004). Examples of antigen-binding proteins include, but are not limited to: human antibodies; humanized antibodies; chimeric antibodies; recombinant antibodies; single-chain antibodies; bifunctional antibodies; trifunctional antibodies; tetrafunctional antibodies; Fab, Fab', Fv fragments, Bs-Fv, F(ab')2, F(ab)2, scFv, di-scFv, dAb, IgD antibodies; IgE antibodies; IgM antibodies; IgG1 antibodies; IgG2 antibodies; IgG3 antibodies; or IgG4 antibodies and fragments thereof.

[0140] In this application, the term "CDR," also known as "complementarity-determining region," typically refers to a region within the variable structural domain of an antibody whose sequence is highly variable and / or forms a structurally defining loop. Typically, antibodies comprise six CDRs: three in the VH (HCDR1, HCDR2, HCDR3) and three in the VL (LCDR1, LCDR2, LCDR3). In some embodiments, naturally occurring camel antibodies consisting only of the heavy chain can function normally and stably even in the absence of the light chain. See, for example, Hamers-Casterman et al., Nature 363:446-448 (1993); Sheriff et al., Nature Struct. Biol. 3:733-736 (1996). Antibody CDRs can be determined using various coding systems, such as CCG, Kabat, AbM, Chothia, IMGT, and a combination of Kabat / Chothia. These coding systems are known in the art, and for example, see http: / / www.bioinf.org.uk / abs / index.html#kabatnum. For example, the amino acid sequence number of the antigen-binding protein can be in accordance with the IMGT numbering scheme (IMGT, the international ImMunoGeneTics informationsystem@imgt.cines.fr; http: / / imgt.cines.fr; Lefranc et al., 1999, Nucleic Acids Res. 27:209-212; Ruiz et al., 2000, Nucleic Acids Res. 28:219-221; Lefranc et al., 2001, Nucleic Acids Res. 29:207-209; Lefranc et al., 2003, Nucleic Acids Res. 31:307-310; Lefranc et al., 2005, DevComp Immunol 29:185-203). For example, the CDR of the antigen-binding protein can be determined according to the Kabat numbering system (see, for example, Kabat EA & Wu TT (1971) Ann NY Acad Sci 190:382-391 and Kabat EA et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, USD Department of Health and Human Services, NIH Publication No. 91-3242).

[0141] In this application, the term "FR" generally refers to the more conserved portion of the antibody variable domain, which is referred to as the frame region. Typically, the variable domains of the natural heavy and light chains each contain four FR regions: four in VH (H-FR1, H-FR2, H-FR3, and H-FR4) and four in VL (L-FR1, L-FR2, L-FR3, and L-FR4).

[0142] In this application, the terms "variable domain" and "variable region" are used interchangeably, generally referring to a portion of the antibody heavy chain and / or light chain. The variable domains of the heavy and light chains can be respectively referred to as "V..." H " and "V L (or referred to as "VH" and "VL" respectively). These domains are usually the most varied parts of the antibody (relative to other antibodies of the same type) and contain antigen-binding sites.

[0143] In this application, the term "variable" generally refers to the fact that certain segments of the variable domain may differ significantly in sequence between antibodies. The variable domain mediates antigen binding and determines the specificity of a particular antibody for its specific antigen. However, variability is not uniformly distributed across the entire variable domain. It is typically concentrated in three segments within the light and heavy chain variable domains, known as hypervariable regions (CDRs or HVRs). The more conserved portions of the variable domain are called framework regions (FRs). The variable domains of the natural heavy and light chains each contain four FR regions, most of which are β-sheet conformations linked by three CDRs forming a ring link, and in some cases forming part of a β-sheet structure. The CDRs in each chain are held together closely by the FR regions, and CDRs from the other chain together promote the formation of the antigen-binding site of the antibody (see Kabat et al, Sequences of Immunological Interest, Fifth Edition, National Institute of Health, Bethesda, MD. (1991)).

[0144] In this application, the term "antibody" generally refers to an immunoglobulin or a fragment thereof or a derivative thereof, encompassing any polypeptide that includes an antigen-binding site, whether it is produced in vitro or in vivo. This term includes, but is not limited to, polyclonal, monoclonal, single-specific, multi-specific, non-specific, humanized, single-chain, chimeric, synthetic, recombinant, hybrid, mutated, and transplanted antibodies. Unless otherwise modified by the term "complete," such as in "complete antibody," for the purposes of this invention, the term "antibody" also includes antibody fragments such as Fab, F(ab')2, Fv, scFv, Fd, dAb, and other antibody fragments that maintain antigen-binding function (e.g., specific binding to Staphylococcus aureus leukocidin). Typically, such fragments should include an antigen-binding domain. The basic 4-chain antibody unit is a heterotetrameric glycoprotein composed of two identical light (L) chains and two identical heavy (H) chains. IgM antibodies consist of five basic heterotetrameric units and a polypeptide chain called the J chain, containing 10 antigen-binding sites. IgA antibodies consist of 2-5 basic tetrameric units that can polymerize with the J chain to form multivalent combinations. For IgG, a tetrameric unit is typically about 150,000 Daltons. Each L chain is linked to an H chain by a covalent disulfide bond, and two H chains are linked to each other by one or more disulfide bonds depending on the H chain isoform. Each H and L chain also has regularly spaced intrachain disulfide bridging bonds. Each H chain has a variable domain (VH) at its N-terminus, followed by three constant domains (CH) for α and γ chains, and four CH domains for μ and ε isoforms. Each L chain has a variable domain (VL) at its N-terminus and a constant domain at its other end. VL corresponds to VH, and CL corresponds to the first constant domain (CH1) of the heavy chain. Specific amino acid residues are thought to form interfaces between the variable domains of the light and heavy chains. VH and VL pair together to form a single antigen-binding site. For the structure and properties of different classes of antibodies, see, for example, Basic and Clinical Immunology, 8th Edition, Daniel P. Sties, Abba I. Terr and Tristram G. Parsolw (eds), Appleton & Lange, Norwalk, Conn., 1994, page 71 and Chapter 6. L chains from any vertebrate species can be classified into one of two distinct types, called κ and λ, based on the amino acid sequence of their constant domains. Immunoglobulins can be classified into different classes or isotypes based on the amino acid sequence of their heavy chain (CH) constant domains. Currently, there are five classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, with heavy chains named α, δ, ε, γ, and μ, respectively.

[0145] In this application, the term "antigen-binding fragment" generally refers to one or more fragments capable of specifically binding antigens (e.g., Staphylococcus aureus leukocidin). In this application, the antigen-binding fragment may include Fab, Fab', F(ab)2, Fv fragments, F(ab')2, scFv, di-scFv, and / or dAb.

[0146] In this application, the term "Fab" generally refers to the antigen-binding fragment of an antibody. As described above, an intact antibody can be digested using papain. After digestion with papain, the antibody produces two identical antigen-binding fragments, namely the "Fab" fragment, and a residual "Fc" fragment (i.e., the Fc region, as above). The Fab fragment may consist of a complete L chain with a variable region of a heavy chain and a first constant region (CH1) of the H chain (VH).

[0147] In this application, the term "Fab' fragment" generally refers to a monovalent antigen-binding fragment of a human monoclonal antibody, which is slightly larger than the Fab fragment. For example, the Fab' fragment may include all light chains, all heavy chain variable regions, and all or part of the first and second constant regions of the heavy chain. For example, the Fab' fragment may also include part or all of the 220-330 amino acid residues of the heavy chain.

[0148] In this application, the term "F(ab')2" generally refers to an antibody fragment produced by digesting an intact antibody with pepsin. The F(ab')2 fragment contains two Fab fragments held together by disulfide bonds and a partial hinge region. The F(ab')2 fragment has bivalent antigen-binding activity and is capable of cross-linking antigens.

[0149] In this application, the term "Fv fragment" generally refers to a monovalent antigen-binding fragment of a human monoclonal antibody, comprising all or part of the heavy chain variable region and light chain variable region, and lacking the heavy chain constant region and light chain constant region. The heavy chain variable region and light chain variable region include, for example, CDRs. For example, an Fv fragment comprises all or part of the amino-terminal variable region of about 110 amino acids of the heavy and light chains.

[0150] In this application, the term "scFv" generally refers to a fusion protein comprising at least one antibody fragment including a variable region comprising a light chain and at least one antibody fragment including a variable region comprising a heavy chain, wherein the light and heavy chain variable regions are adjacent (e.g., via a synthetic linker, such as a short, flexible peptide linker) and are capable of being expressed as a single-chain polypeptide, and wherein the scFv retains the specificity of the intact antibody from which it is derived. Unless otherwise specified, as used in this application, the scFv may have the VL and VH variable regions in any order (e.g., relative to the N-terminus and C-terminus of the polypeptide), and the scFv may comprise VL-linker-VH or may comprise VH-linker-VL.

[0151] In this application, the term "dAb" generally refers to an antigen-binding fragment having a VH domain, a VL domain, or having either a VH domain or a VL domain, for example, Ward et al. (Nature, 1989 Oct 12; 341(6242): 544-6), Holt et al., Trends Biotechnol., 2003, 21(11): 484-490; and other published patent applications, such as WO 06 / 030220, WO 06 / 003388, and Domantis Ltd. The term "dAb" generally includes sdAb. The term "sdAb" generally refers to a single-domain antibody. A single-domain antibody generally refers to an antibody fragment consisting only of the variable region (VH domain) of the antibody heavy chain or the variable region (VL) of the antibody light chain.

[0152] In this application, the term "monoclonal antibody" generally refers to an antibody molecule preparation consisting of a single molecule. Monoclonal antibodies typically exhibit high specificity against a single antigenic site. Moreover, unlike conventional polyclonal antibody preparations (which usually contain different antibodies targeting different determinants), each monoclonal antibody targets a single determinant on the antigen. In addition to their specificity, monoclonal antibodies have the advantage that they can be synthesized through hybridoma culture without contamination by other immunoglobulins. The modifier "monoclonal" indicates the characteristic of antibodies obtained from a substantially homogeneous group of antibodies and is not to be interpreted as requiring the production of antibodies through any particular method. For example, the monoclonal antibodies used in this application can be prepared in hybridoma cells or through recombinant DNA methods.

[0153] In this application, the term "chimeric antibody" generally refers to an antibody in which the variable region is derived from one species and the constant region is derived from another species. Typically, the variable region is derived from an antibody from a laboratory animal such as a rodent ("parental antibody") and the constant region is derived from a human antibody, such that the resulting chimeric antibody is less likely to induce an adverse immune response in human individuals compared to parental (e.g., alpaca-derived) antibodies.

[0154] In this application, the term "humanized antibody" generally refers to an antibody in which some or all of the amino acids outside the CDR region of a non-human antibody (e.g., an alpaca antibody) are replaced by corresponding amino acids derived from human immunoglobulins. Small additions, deletions, insertions, substitutions, or modifications of amino acids within the CDR region are also permissible, as long as they retain the antibody's ability to bind to a specific antigen. Humanized antibodies may optionally contain at least a portion of the constant region of human immunoglobulins. "Humanized antibodies" retain antigen specificity similar to the original antibody. The "humanized" form of a non-human (e.g., alpaca) antibody may minimally contain a chimeric antibody with a sequence derived from a non-human immunoglobulin. In some cases, CDR region residues in a human immunoglobulin (receptor antibody) may be replaced with CDR region residues from a non-human species (donor antibody) (such as an alpaca, mouse, rat, rabbit, or non-human primate) having the desired properties, affinity, and / or capabilities. In some cases, FR region residues in a human immunoglobulin may be replaced with corresponding non-human residues. Furthermore, humanized antibodies may contain amino acid modifications not found in receptor antibodies or in donor antibodies. These modifications can be made to further improve antibody performance, such as binding affinity.

[0155] The term "fully human antibody" generally refers to an antibody that contains only the sequence of human immunoglobulin proteins. If it is produced in mice, in mouse cells, or in hybridomas derived from mouse cells, then a fully human antibody may contain mouse sugar chains. Similarly, "mouse antibody" or "rat antibody" refers to an antibody that contains only the sequence of mouse or rat immunoglobulins, respectively. Fully human antibodies can be generated in humans or in transgenic animals with germline sequences of human immunoglobulins by phage display or other molecular biology methods. Exemplary techniques that can be used to manufacture antibodies are described in U.S. Patents: 6,150,584, 6,458,592, and 6,420,140. Other techniques, such as the use of libraries, are known in the art.

[0156] In this application, the term "cross-neutralization" generally refers to the neutralization of two or more toxins. The toxins may include leukocytoxins. For example, the leukocytoxins may include hemolysin and / or PVL leukocytoxin. For example, the hemolysin may include gamma-hemolysin.

[0157] In this application, the terms "peptide molecule," "polypeptide," and "peptide" are used interchangeably and generally refer to polymers of amino acid residues. The term "fusion protein" generally refers to a polypeptide having at least two parts covalently linked together. Each part can be a polypeptide with different properties. These properties can be biological properties, such as in vitro or in vivo activity. They can also be simple chemical or physical properties, such as binding to target molecules, catalysis of reactions, etc. The two parts can be linked by a single peptide bond or through a peptide linker.

[0158] In this application, the term "nucleic acid molecule" generally refers to a nucleotide, deoxyribonucleotide, or ribonucleotide of any length in an isolated form, or an analogue isolated from its natural environment or synthesized artificially.

[0159] In this application, the term "vector" generally refers to a nucleic acid delivery vehicle that can insert a polynucleotide encoding a protein into itself and enable the protein to be expressed. Vectors can transform, transduce, or transfect host cells, allowing the genetic material elements they carry to be expressed within the host cells. For example, vectors can include: plasmids; phage particles; Cos plasmids; artificial chromosomes such as yeast artificial chromosomes (YAC), bacterial artificial chromosomes (BAC), or P1-derived artificial chromosomes (PAC); bacteriophages such as λ phage or M13 phage; and animal viruses. Animal viruses used as vectors can include retroviruses (including lentiviruses), adenoviruses, adeno-associated viruses, herpesviruses (such as herpes simplex virus), poxviruses, baculoviruses, papillomaviruses, and papillomaviruses (such as SV40). A vector may contain multiple elements controlling expression, including promoter sequences, transcription initiation sequences, enhancer sequences, selection elements, and reporter genes. Additionally, vectors may contain a replication initiation site. The carrier may also include components that help it enter the cell, such as viral particles, liposomes, or protein coats, but not only these substances.

[0160] In this application, the term "cell" generally refers to a single cell, cell line, or cell culture that may be or is already a recipient of a subject plasmid or vector, including the nucleic acid molecules or vectors described in this invention. Cells may include the progeny of a single cell. Due to natural, accidental, or intentional mutations, the progeny may not necessarily be identical to the original parent cell (in terms of the morphology of the total DNA complement or in the genome). Cells may include cells transfected in vitro using the vectors described in this application. Cells may be bacterial cells (e.g., *E. coli*), yeast cells, or other eukaryotic cells, such as COS cells, Chinese hamster ovary (CHO) cells, CHO-K1 cells, LNCAP cells, HeLa cells, HEK293 cells, COS-1 cells, and NSO cells.

[0161] In this application, the term "immunoconjugate" generally refers to a conjugate formed by conjugating the other reagents (e.g., chemotherapeutic agents, radioactive elements, cell growth inhibitors, and cytotoxic agents) with the antibody or its antigen-binding fragment (e.g., covalently linked by a linker molecule), which can deliver the other reagents to the target cells by specifically binding the antibody or its antigen-binding fragment to the antigen on the target cells.

[0162] In this application, the term "pharmaceutical composition" generally refers to a composition for the prevention / treatment of a disease or condition. The pharmaceutical composition may comprise the isolated antigen-binding protein described in this application, the nucleic acid molecule described in this application, the carrier described in this application, and / or the cell described in this application, and optionally a pharmaceutically acceptable adjuvant. Furthermore, the pharmaceutical composition may also comprise suitable formulations of one or more (pharmaceutically effective) carriers, stabilizers, excipients, diluents, solubilizers, surfactants, emulsifiers, and / or preservatives. The acceptable components of the composition are preferably non-toxic to the recipient at the dosage and concentration used. The pharmaceutical compositions of the present invention include, but are not limited to, liquid, freeze-dried, and lyophilized compositions.

[0163] In this application, the term "pharmaceutically acceptable carrier" generally includes pharmaceutically acceptable carriers, excipients, or stabilizers that are non-toxic to cells or mammals exposed to them at the doses and concentrations employed. Physiologically acceptable carriers may include, for example, buffers, antioxidants, low molecular weight (less than about 10 residues) peptides, proteins, hydrophilic polymers, amino acids, monosaccharides, disaccharides and other carbohydrates, chelating agents, sugar alcohols, salt-forming anti-charge ions such as sodium, and / or nonionic surfactants.

[0164] In this application, the terms "specific binding" or "specific" generally refer to measurable and reproducible interactions, such as binding between a target and an antibody, where the presence of the target is determined by the presence of a heterogeneous population of molecules (including biomolecules). For example, an antibody that specifically binds to a target (which may be an epitope) may be an antibody that binds to the target with greater affinity, affinity, ease, and / or a longer duration than it binds to other targets. In some embodiments, the antibody specifically binds to an epitope on a protein that is conserved across proteins of different species. In some embodiments, specific binding may include, but is not required to be, exclusive binding. The term also applies to antigen-binding proteins that are specific to a particular epitope that cross-reacts with multiple antigens, wherein a specific antibody can bind to multiple antigens carrying cross-reactive epitopes. Such binding sites of antigen-binding proteins and / or antigen-binding proteins having specific binding to cross-reactive epitopes are also referred to as multispecific or cross-specific binding sites, respectively. For example, an antigen-binding protein may have a multispecific binding site that specifically binds to epitopes that cross-react with multiple different antigens.

[0165] In this application, the term "subject" generally refers to a human or non-human animal, including but not limited to cats, dogs, horses, pigs, cows, sheep, rabbits, mice, rats, or monkeys.

[0166] In this application, the proteins, peptides and / or amino acid sequences involved should also be understood to include at least the following range: variants or homologs that have the same or similar functions as the said protein or peptide.

[0167] In this application, the variant can be, for example, a protein or polypeptide that has undergone substitution, deletion, or addition of one or more amino acids in the amino acid sequence of the protein and / or the polypeptide (e.g., an antibody that specifically binds to Staphylococcus aureus leukocidin or a fragment thereof). For example, the functional variant may comprise a protein or polypeptide that has undergone amino acid alterations through substitution, deletion, and / or insertion of at least one, such as 1-30, 1-20, or 1-10, or even 1, 2, 3, 4, or 5 amino acids. The functional variant may substantially retain the biological properties of the protein or polypeptide prior to the alteration (e.g., substitution, deletion, or addition). For example, the functional variant may retain at least 60%, 70%, 80%, 90%, or 100% of the biological activity (e.g., antigen-binding capacity) of the protein or polypeptide prior to the alteration. For example, the substitution may be a conserved substitution.

[0168] In this application, the homolog can be a protein or polypeptide having at least about 85% (e.g., having at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99% or higher) sequence homology with the amino acid sequence of the protein and / or the polypeptide (e.g., an antibody that specifically binds to Staphylococcus aureus leukocidin or a fragment thereof).

[0169] In this application, homology generally refers to the similarity, resemblance, or association between two or more sequences. The "sequence homology percentage" can be calculated by comparing two sequences to be aligned within a comparison window, determining the number of positions in the two sequences containing the same nucleic acid bases (e.g., A, T, C, G, I) or the same amino acid residues (e.g., Ala, Pro, Ser, Thr, Gly, Val, Leu, Ile, Phe, Tyr, Trp, Lys, Arg, His, Asp, Glu, Asn, Gln, Cys, and Met) to obtain the number of matching positions, dividing the number of matching positions by the total number of positions in the comparison window (i.e., the window size), and multiplying the result by 100 to produce the sequence homology percentage. Alignments performed to determine the sequence homology percentage can be performed in various ways known in the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine suitable parameters for aligning sequences, including any algorithm required to achieve maximum alignment across the full-length sequence being compared or within the target sequence region. Homology can also be determined using FASTA and BLAST. A description of the FASTA algorithm can be found in W.R. Pearson and DJ Lipman, “An Improved Tool for Biological Sequence Alignment,” Proceedings of the National Academy of Sciences (Proc. Natl. Acad. Sci.), 85: 2444-2448, 1988; and DJ Lipman and W.R. Pearson, “A Fast and Sensitive Search for Protein Similarity,” Science, 227: 1435-1441, 1989. A description of the BLAST algorithm can be found in S. Altschul, W. Gish, W. Miller, E.W. Myers, and D. Lipman, “A Basic Tool for Local Alignment Search,” Journal of Molecular Biology, 215: 403-410, 1990.

[0170] In this application, the term "comprising" generally means including, encompassing, containing, or including. In some cases, it also means "to be" or "composed of".

[0171] In this application, the term "about" generally refers to a variation within a range of 0.5% to 10% above or below a specified value, such as a variation within a range of 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, 5.5%, 6%, 6.5%, 7%, 7.5%, 8%, 8.5%, 9%, 9.5%, or 10% above or below a specified value. Invention Details

[0173] Isolated antigen-binding proteins

[0174] The CDR (Complementarity Determinant Region) of an antibody, also known as the complementarity-determining region, is part of the variable region. Amino acid residues in this region can contact antigens or antigenic epitopes. Antibody CDRs can be determined using various coding systems, such as CCG, Kabat, Chothia, IMGT, AbM, and a combination of Kabat / Chothia. These coding systems are known in the art and can be found, for example, at http: / / www.bioinf.org.uk / abs / index.html#kabatnum. Those skilled in the art can determine the CDR region using different coding systems based on the antibody's sequence and structure. The CDR region may differ when using different coding systems. In this application, the term CDR encompasses CDR sequences partitioned according to any CDR partitioning method; it also encompasses variants of the CDR, including amino acid sequences with substitutions, deletions, and / or additions of one or more amino acids. For example, 1-30, 1-20, or 1-10 amino acid substitutions, deletions, and / or insertions, or 1, 2, 3, 4, 5, 6, 7, 8, or 9 amino acid substitutions, deletions, and / or insertions; homologs are also included, which can be amino acid sequences having at least about 85% (e.g., having at least about 85%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98%, about 99%, or higher) sequence homology with the amino acid sequence of the CDR. In some embodiments, the isolated antigen-binding protein described in this application is defined by the Kabat coding system.

[0175] On one hand, this application provides an isolated antigen-binding protein that may contain at least one CDR in the variable region (VH) of the antibody heavy chain, wherein the VH may contain the amino acid sequence shown in SEQ ID NO:53. For example, the VH may contain the amino acid sequence shown in any one of SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, and SEQ ID NO:35. In this application, the HCDR of the isolated antigen-binding protein can be divided in any form, as long as the VH is identical to the amino acid sequence shown in any one of SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, and SEQ ID NO:35, and the HCDR obtained by any form of division falls within the protection scope of this application.

[0176] In this application, the isolated antigen-binding protein may contain HCDR3, which may contain the amino acid sequence shown in SEQ ID NO:1.

[0177] In this application, the isolated antigen-binding protein may contain HCDR2, which may contain the amino acid sequence shown in SEQ ID NO:2.

[0178] In this application, the isolated antigen-binding protein may include HCDR1, which may contain the amino acid sequence shown in SEQ ID NO:3.

[0179] In this application, the isolated antigen-binding protein may include HCDR1, HCDR2 and HCDR3, wherein HCDR1 may include the amino acid sequence shown in SEQ ID NO:3, HCDR2 may include the amino acid sequence shown in SEQ ID NO:2 and HCDR3 may include the amino acid sequence shown in SEQ ID NO:1.

[0180] In this application, the isolated antigen-binding protein may include H-FR1, the C-terminus of which is directly or indirectly linked to the N-terminus of HCDR1, and the H-FR1 may include the amino acid sequence shown in SEQ ID NO:45.

[0181] In this application, H-FR1 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:10. For example, the substitution may be at one or more positions selected from the group consisting of: position 1, position 5, position 11, position 12, position 13, position 14, position 20, and position 23. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:10. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:14. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:18.

[0182] In this application, the isolated antigen-binding protein may include H-FR2, which is located between HCDR1 and HCDR2, and H-FR2 may include the amino acid sequence shown in SEQ ID NO:46.

[0183] In this application, H-FR2 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:11. For example, the substitution may be at one or more positions selected from the group consisting of: position 3, position 5, position 7, and position 13. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:11. For example, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:15. For example, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:19.

[0184] In this application, the isolated antigen-binding protein may include H-FR3, which is located between HCDR2 and HCDR3, and the H-FR3 may include the amino acid sequence shown in SEQ ID NO:47.

[0185] In this application, H-FR3 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:12. For example, the substitution may be at one or more positions selected from the group consisting of: position 1, position 2, position 10, position 11, position 15, position 16, position 19, position 21, position 23, and position 29. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:12. For example, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:16. For example, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:20.

[0186] In this application, the isolated antigen-binding protein may include H-FR4, the N-terminus of which is linked to the C-terminus of HCDR3, and the H-FR4 contains the amino acid sequence shown in SEQ ID NO:48.

[0187] In this application, H-FR4 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:13. For example, the substitution may be at one or more positions selected from the group consisting of positions 6 and 11. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, H-FR4 may contain the amino acid sequence shown in SEQ ID NO:13. For example, H-FR4 may contain the amino acid sequence shown in SEQ ID NO:17.

[0188] In this application, the isolated antigen-binding protein may comprise H-FR1, H-FR2, H-FR3, and H-FR4. H-FR1 may comprise the amino acid sequence shown in SEQ ID NO:45, H-FR2 may comprise the amino acid sequence shown in SEQ ID NO:46, H-FR3 may comprise the amino acid sequence shown in SEQ ID NO:47, and H-FR4 may comprise the amino acid sequence shown in SEQ ID NO:48. For example, H-FR1 may comprise the amino acid sequence shown in any one of SEQ ID NO:10, SEQ ID NO:14, and SEQ ID NO:18; H-FR2 may comprise the amino acid sequence shown in any one of SEQ ID NO:11, SEQ ID NO:15, and SEQ ID NO:19; H-FR3 may comprise the amino acid sequence shown in any one of SEQ ID NO:12, SEQ ID NO:16, and SEQ ID NO:20; and H-FR4 may comprise the amino acid sequence shown in SEQ ID NO:13 or SEQ ID NO:17. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:10, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:11, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:12, and H-FR4 may contain the amino acid sequence shown in SEQ ID NO:13. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:14, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:15, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:16, and H-FR4 may contain the amino acid sequence shown in SEQ ID NO:17. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:18, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:15, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:16, and H-FR4 may contain the amino acid sequence shown in SEQ ID NO:17. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:18, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:19, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:16, and H-FR4 may contain the amino acid sequence shown in SEQ ID NO:17.For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:18, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:15, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:20, and H-FR4 may contain the amino acid sequence shown in SEQ ID NO:17. For example, H-FR1 may contain the amino acid sequence shown in SEQ ID NO:18, H-FR2 may contain the amino acid sequence shown in SEQ ID NO:19, H-FR3 may contain the amino acid sequence shown in SEQ ID NO:20, and H-FR4 may contain the amino acid sequence shown in SEQ ID NO:17.

[0189] In this application, the isolated antigen-binding protein may comprise HCDR1, HCDR2, HCDR3, H-FR1, H-FR2, H-FR3, and H-FR4. For example, the isolated antigen-binding protein may comprise the amino acid sequences shown in SEQ ID NO:3, SEQ ID NO:2, SEQ ID NO:1, SEQ ID NO:10, SEQ ID NO:11, SEQ ID NO:12, and SEQ ID NO:13, respectively. For example, the isolated antigen-binding protein may comprise the amino acid sequences shown in SEQ ID NO:3, SEQ ID NO:2, SEQ ID NO:1, SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, and SEQ ID NO:17, respectively. For example, the isolated antigen-binding proteins HCDR1, HCDR2, HCDR3, H-FR1, H-FR2, H-FR3, and H-FR4 may, respectively, contain the amino acid sequences shown in SEQ ID NO:3, SEQ ID NO:2, SEQ ID NO:1, SEQ ID NO:18, SEQ ID NO:15, SEQ ID NO:16, and SEQ ID NO:17. For example, the isolated antigen-binding proteins HCDR1, HCDR2, HCDR3, H-FR1, H-FR2, H-FR3, and H-FR4 may, respectively, contain the amino acid sequences shown in SEQ ID NO:3, SEQ ID NO:2, SEQ ID NO:1, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:16, and SEQ ID NO:17. For example, the isolated antigen-binding proteins HCDR1, HCDR2, HCDR3, H-FR1, H-FR2, H-FR3 and H-FR4 may respectively contain the amino acid sequences shown in SEQ ID NO:3, SEQ ID NO:2, SEQ ID NO:1, SEQ ID NO:18, SEQ ID NO:15, SEQ ID NO:20 and SEQ ID NO:17.For example, the isolated antigen-binding proteins HCDR1, HCDR2, HCDR3, H-FR1, H-FR2, H-FR3 and H-FR4 may respectively contain the amino acid sequences shown in SEQ ID NO:3, SEQ ID NO:2, SEQ ID NO:1, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:20 and SEQ ID NO:17.

[0190] In this application, the isolated antigen-binding protein may contain VH, which contains the amino acid sequence shown in SEQ ID NO:53.

[0191] In this application, the VH may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:30. For example, the substitution may be at one or more positions selected from the group consisting of: position 1, position 5, position 11, position 12, position 13, position 14, position 20, position 23, position 38, position 40, position 42, position 48, position 67, position 68, position 76, position 77, position 81, position 82, position 85, position 87, position 89, position 95, position 112, and position 117. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, the VH may contain the amino acid sequence shown in any one of SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, and SEQ ID NO:35.

[0192] In this application, the isolated antigen-binding protein may include an antibody heavy chain constant region. For example, the antibody heavy chain constant region may be derived from the heavy chain constant region of any immunoglobulin, including IgM, IgD, IgG, IgA, and IgE. For example, the antibody heavy chain constant region may be derived from the human IgG heavy chain constant region. In this application, the immunoglobulin heavy chain constant region may contain mutants thereof. In this application, the antibody heavy chain constant region may be derived from the heavy chain constant region of any one of human IgG1-4. For example, the isolated antigen-binding protein may be derived from the human IgG1 heavy chain constant region. For example, the heavy chain constant region may contain the amino acid sequence shown in SEQ ID NO:42.

[0193] In this application, the isolated antigen-binding protein may include a heavy chain, which may include a VH (volume-highest amino acid) and a heavy chain constant region. For example, the VH of the heavy chain may include the amino acid sequence shown in SEQ ID NO:30, and the heavy chain constant region may include the amino acid sequence shown in SEQ ID NO:42. For example, the VH of the heavy chain may include the amino acid sequence shown in SEQ ID NO:31, and the heavy chain constant region may include the amino acid sequence shown in SEQ ID NO:42. For example, the VH of the heavy chain may include the amino acid sequence shown in SEQ ID NO:32, and the heavy chain constant region may include the amino acid sequence shown in SEQ ID NO:42. For example, the VH of the heavy chain may include the amino acid sequence shown in SEQ ID NO:33, and the heavy chain constant region may include the amino acid sequence shown in SEQ ID NO:42. For example, the VH of the heavy chain may include the amino acid sequence shown in SEQ ID NO:34, and the heavy chain constant region may include the amino acid sequence shown in SEQ ID NO:42. For example, the VH of the heavy chain may include the amino acid sequence shown in SEQ ID NO:35, and the heavy chain constant region may include the amino acid sequence shown in SEQ ID NO:42. For example, the heavy chain may include the amino acid sequence shown in any one of SEQ ID NO:55, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60 and SEQ ID NO:61.

[0194] In this application, the isolated antigen-binding protein may contain at least one CDR in the variable region (VL) of the antibody light chain, and the VL may contain the amino acid sequence shown in SEQ ID NO:54. For example, the VL may contain the amino acid sequence shown in any one of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, and SEQ ID NO:41. In this application, the LCDR of the isolated antigen-binding protein can be divided in any form, as long as the VL is identical to the amino acid sequence shown in any one of SEQ ID NO:54, SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, and SEQ ID NO:41. The LCDR obtained by dividing in any form falls within the protection scope of this application.

[0195] In this application, the isolated antigen-binding protein may include LCDR3, which may contain the amino acid sequence shown in SEQ ID NO:4.

[0196] In this application, the isolated antigen-binding protein may include LCDR2, which may contain the amino acid sequence shown in SEQ ID NO:5.

[0197] In this application, the isolated antigen-binding protein may include LCDR1, which may contain the amino acid sequence shown in SEQ ID NO:44.

[0198] In this application, the L-CDR1 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:6. For example, the substitution may be at one or more positions selected from the group consisting of: position 8, position 11, and position 12. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, the LCDR1 may contain the amino acid sequence shown in SEQ ID NO:6. For example, the LCDR1 may contain the amino acid sequence shown in SEQ ID NO:7. For example, the LCDR1 may contain the amino acid sequence shown in SEQ ID NO:8. For example, the LCDR1 may contain the amino acid sequence shown in SEQ ID NO:9.

[0199] In this application, the isolated antigen-binding protein may comprise LCDR1, LCDR2, and LCDR3, wherein LCDR1 may comprise the amino acid sequence shown in SEQ ID NO:44, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4.

[0200] In this application, the isolated antigen-binding protein may comprise LCDR1, LCDR2, and LCDR3. For example, LCDR1 may comprise the amino acid sequence shown in SEQ ID NO:6, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4. For example, LCDR1 may comprise the amino acid sequence shown in SEQ ID NO:7, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4. For example, LCDR1 may comprise the amino acid sequence shown in SEQ ID NO:8, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4. For example, LCDR1 may comprise the amino acid sequence shown in SEQ ID NO:9, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4.

[0201] In this application, the isolated antigen-binding protein may include L-FR1, the C-terminus of which is directly or indirectly linked to the N-terminus of LCDR1, and the L-FR1 may include the amino acid sequence shown in SEQ ID NO:49.

[0202] In this application, L-FR1 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:21. For example, the substitution may be at one or more positions selected from the group consisting of: position 2, position 11, position 14, position 17, position 14, and position 18. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, L-FR1 may contain the amino acid sequence shown in SEQ ID NO:21. For example, L-FR1 may contain the amino acid sequence shown in SEQ ID NO:25. For example, L-FR1 may contain the amino acid sequence shown in SEQ ID NO:29.

[0203] In this application, the isolated antigen-binding protein may include L-FR2, which is located between LCDR1 and LCDR2, and L-FR2 may include the amino acid sequence shown in SEQ ID NO:50.

[0204] In this application, L-FR2 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:22. For example, the substitution may be at one or more positions selected from the group consisting of: position 3, position 5, position 9, and position 11. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be the replacement of an amino acid residue with an amino acid residue having a similar side chain. For example, L-FR2 may contain the amino acid sequence shown in SEQ ID NO:22. For example, L-FR2 may contain the amino acid sequence shown in SEQ ID NO:26.

[0205] In this application, the isolated antigen-binding protein may include L-FR3, which is located between LCDR2 and LCDR3, and L-FR3 may include the amino acid sequence shown in SEQ ID NO:51.

[0206] In this application, the L-FR3 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:23. For example, the substitution may be at one or more positions selected from the group consisting of: position 11, position 12, position 27, and position 31. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be the replacement of an amino acid residue with an amino acid residue having a similar side chain. For example, the L-FR3 may contain the amino acid sequence shown in SEQ ID NO:23. For example, the L-FR3 may contain the amino acid sequence shown in SEQ ID NO:27.

[0207] In this application, the isolated antigen-binding protein may include L-FR4, the N-terminus of which is connected to the C-terminus of LCDR3, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:52.

[0208] In this application, the L-FR4 may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:24. For example, the substitution may be at position 3. For example, the substitution may be a conservative substitution of the amino acid. For example, the conservative substitution may be replacing an amino acid residue with an amino acid residue having a similar side chain. For example, the L-FR4 may contain the amino acid sequence shown in SEQ ID NO:24. For example, the L-FR4 may contain the amino acid sequence shown in SEQ ID NO:28.

[0209] In this application, the isolated antigen-binding protein may comprise L-FR1, L-FR2, L-FR3, and L-FR4. L-FR1 may comprise the amino acid sequence shown in SEQ ID NO:49, L-FR2 may comprise the amino acid sequence shown in SEQ ID NO:50, L-FR3 may comprise the amino acid sequence shown in SEQ ID NO:51, and L-FR4 may comprise the amino acid sequence shown in SEQ ID NO:52. For example, L-FR1 may comprise the amino acid sequence shown in any one of SEQ ID NO:21, SEQ ID NO:25, and SEQ ID NO:29; L-FR2 may comprise the amino acid sequence shown in SEQ ID NO:22 or SEQ ID NO:26; L-FR3 may comprise the amino acid sequence shown in SEQ ID NO:23 or SEQ ID NO:27; and L-FR4 may comprise the amino acid sequence shown in SEQ ID NO:24 or SEQ ID NO:28. For example, L-FR1 may contain the amino acid sequence shown in SEQ ID NO:21, L-FR2 may contain the amino acid sequence shown in SEQ ID NO:22, L-FR3 may contain the amino acid sequence shown in SEQ ID NO:23, and L-FR4 may contain the amino acid sequence shown in SEQ ID NO:24; for example, L-FR1 may contain the amino acid sequence shown in SEQ ID NO:25, L-FR2 may contain the amino acid sequence shown in SEQ ID NO:26, L-FR3 may contain the amino acid sequence shown in SEQ ID NO:27, and L-FR4 may contain the amino acid sequence shown in SEQ ID NO:28; for example, L-FR1 may contain the amino acid sequence shown in SEQ ID NO:29, L-FR2 may contain the amino acid sequence shown in SEQ ID NO:26, L-FR3 may contain the amino acid sequence shown in SEQ ID NO:27, and L-FR4 may contain the amino acid sequence shown in SEQ ID NO:28.

[0210] In this application, the isolated antigen-binding protein may comprise LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3, and L-FR4. For example, the isolated antigen-binding protein's LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3, and L-FR4 may respectively comprise the amino acid sequences shown in SEQ ID NO:6, SEQ ID NO:5, SEQ ID NO:4, SEQ ID NO:21, SEQ ID NO:22, SEQ ID NO:23, and SEQ ID NO:24. For example, the isolated antigen-binding protein's LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3, and L-FR4 may respectively comprise the amino acid sequences shown in SEQ ID NO:7, SEQ ID NO:5, SEQ ID NO:4, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, and SEQ ID NO:28. For example, the isolated antigen-binding proteins LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3, and L-FR4 may, respectively, contain the amino acid sequences shown in SEQ ID NO:8, SEQ ID NO:5, SEQ ID NO:4, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, and SEQ ID NO:28. For example, the isolated antigen-binding proteins LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3, and L-FR4 may, respectively, contain the amino acid sequences shown in SEQ ID NO:9, SEQ ID NO:5, SEQ ID NO:4, SEQ ID NO:25, SEQ ID NO:26, SEQ ID NO:27, and SEQ ID NO:28. For example, the isolated antigen-binding proteins LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3 and L-FR4 may respectively contain the amino acid sequences shown in SEQ ID NO:8, SEQ ID NO:5, SEQ ID NO:4, SEQ ID NO:29, SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28.For example, the isolated antigen-binding proteins LCDR1, LCDR2, LCDR3, L-FR1, L-FR2, L-FR3 and L-FR4 may respectively contain the amino acid sequences shown in SEQ ID NO:9, SEQ ID NO:5, SEQ ID NO:4, SEQ ID NO:29, SEQ ID NO:26, SEQ ID NO:27 and SEQ ID NO:28.

[0211] In this application, the isolated antigen-binding protein may contain a VL, which contains the amino acid sequence shown in SEQ ID NO:54.

[0212] In this application, the VL may contain one or more amino acid substitutions compared to the amino acid sequence shown in SEQ ID NO:36. For example, the substitution may be at one or more positions selected from the group consisting of: position 2, position 11, position 14, position 17, position 18, position 31, position 34, position 35, position 42, position 44, position 48, position 50, position 72, position 88, position 92, and position 115. For example, the VL may contain the amino acid sequence shown in SEQ ID NO:36. For example, the VL may contain the amino acid sequence shown in SEQ ID NO:37. For example, the VL may contain the amino acid sequence shown in SEQ ID NO:38. For example, the VL may contain the amino acid sequence shown in SEQ ID NO:39. For example, the VL may contain the amino acid sequence shown in SEQ ID NO:40. For example, the VL may contain the amino acid sequence shown in SEQ ID NO:41.

[0213] In this application, the isolated antigen-binding protein may comprise an antibody light chain constant region. For example, the light chain constant region may be derived from a human antibody light chain constant region. For example, the light chain constant region may be derived from a human Igκ constant region. For example, the light chain constant region may comprise the amino acid sequence shown in SEQ ID NO:43.

[0214] In this application, the isolated antigen-binding protein may include a light chain, which may include a volume chain volume (VL) and a light chain constant region. For example, the VL of the light chain may contain the amino acid sequence shown in SEQ ID NO:36, and the light chain constant region may contain the amino acid sequence shown in SEQ ID NO:43. For example, the VL of the light chain may contain the amino acid sequence shown in SEQ ID NO:37, and the light chain constant region may contain the amino acid sequence shown in SEQ ID NO:43. For example, the VL of the light chain may contain the amino acid sequence shown in SEQ ID NO:38, and the light chain constant region may contain the amino acid sequence shown in SEQ ID NO:43. For example, the VL of the light chain may contain the amino acid sequence shown in SEQ ID NO:39, and the light chain constant region may contain the amino acid sequence shown in SEQ ID NO:43. For example, the VL of the light chain may contain the amino acid sequence shown in SEQ ID NO:40, and the light chain constant region may contain the amino acid sequence shown in SEQ ID NO:43. For example, the VL of the light chain may contain the amino acid sequence shown in SEQ ID NO:41, and the constant region of the light chain may contain the amino acid sequence shown in SEQ ID NO:43. For example, the light chain may include the amino acid sequence shown in any one of SEQ ID NO:56, SEQ ID NO:62, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:65, and SEQ ID NO:66.

[0215] In this application, the isolated antigen-binding protein may comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3. For example, HCDR1 of the isolated antigen-binding protein may comprise the amino acid sequence shown in SEQ ID NO:3, HCDR2 may comprise the amino acid sequence shown in SEQ ID NO:2, HCDR3 may comprise the amino acid sequence shown in SEQ ID NO:1, LCDR1 may comprise the amino acid sequence shown in SEQ ID NO:44, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4.

[0216] In this application, the isolated antigen-binding protein may comprise HCDR1, HCDR2, HCDR3, LCDR1, LCDR2, and LCDR3. For example, HCDR1 of the isolated antigen-binding protein may comprise the amino acid sequence shown in SEQ ID NO:3, HCDR2 may comprise the amino acid sequence shown in SEQ ID NO:2, HCDR3 may comprise the amino acid sequence shown in SEQ ID NO:1, LCDR1 may comprise the amino acid sequence shown in any one of SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, and SEQ ID NO:9, LCDR2 may comprise the amino acid sequence shown in SEQ ID NO:5, and LCDR3 may comprise the amino acid sequence shown in SEQ ID NO:4.

[0217] In this application, the isolated antigen-binding protein may comprise VH and VL. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:53, and the VL may comprise the amino acid sequence shown in SEQ ID NO:54. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:30, and the VL may comprise the amino acid sequence shown in SEQ ID NO:36. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:30, and the VL may comprise the amino acid sequence shown in SEQ ID NO:37. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:30, and the VL may comprise the amino acid sequence shown in SEQ ID NO:38. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:30, and the VL may comprise the amino acid sequence shown in SEQ ID NO:39. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:30, and the VL may comprise the amino acid sequence shown in SEQ ID NO:40. For example, the VH may comprise the amino acid sequence shown in SEQ ID NO:30, and the VL may comprise the amino acid sequence shown in SEQ ID NO:41. For example, the VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:36. For example, the VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:37. For example, the VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:38. For example, the VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:39. For example, the VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:40. For example, the VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:41. For example, the VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:36. For example, the VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:37. Alternatively, the VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:38.For example, the VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:39. For example, the VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:40. For example, the VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:41. For example, the VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:36. For example, the VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:37. For example, the VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:38. For example, the VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:39. For example, the VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:40. For example, the VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:41. For example, the VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:36. For example, the VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:37. For example, the VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:38. For example, the VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:39. For example, the VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:40. For example, the VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:41. For example, the VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:36. For example, the VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:37.For example, the VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:38. For example, the VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:39. For example, the VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:40. For example, the VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:41.

[0218] In this application, the isolated antigen-binding protein may comprise a VH, a heavy chain constant region, a VL, and a light chain constant region. For example, the VH may be selected from the amino acid sequence shown in any one of SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, and SEQ ID NO:35; the heavy chain constant region may comprise the amino acid sequence shown in SEQ ID NO:42; the VL may be selected from the amino acid sequence shown in any one of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, and SEQ ID NO:41; and the light chain constant region may comprise the amino acid sequence shown in SEQ ID NO:43.

[0219] In this application, the isolated antigen-binding protein may comprise an antibody or an antigen-binding fragment thereof. In this application, the antigen-binding fragment may include Fab, Fab', Fv fragments, F(ab')2, F(ab)2, scFv, di-scFv, and / or dAb. In this application, the antibody may include monoclonal antibodies, chimeric antibodies, humanized antibodies, and / or fully human antibodies. For example, the VH of the humanized antigen-binding protein may comprise the amino acid sequence shown in any one of SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34, and SEQ ID NO:35. For example, the VL of the humanized antigen-binding protein may comprise the amino acid sequence shown in any one of SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40, and SEQ ID NO:41.

[0220] Furthermore, it should be noted that the isolated antigen-binding protein described in this application may contain heavy chain and / or light chain sequences with one or more conserved sequence modifications. "Conserved sequence modification" refers to amino acid modifications that do not significantly affect or alter antibody binding properties. Such conserved modifications include amino acid substitutions, additions, and deletions. Modifications can be introduced into the isolated antigen-binding protein described in this application using standard techniques known in the art, such as point mutations and PCR-mediated mutations. Conserved amino acid substitution involves replacing an amino acid residue with an amino acid residue having a similar side chain. Groups of amino acid residues with similar side chains are known in the art. These amino acid residue groups include amino acids with basic side chains (e.g., lysine, arginine, histidine), acidic side chains (e.g., aspartic acid, glutamic acid), nonpolar side chains (e.g., glycine, asparagine, glutamine, serine, threonine, tyrosine, cysteine, tryptophan), nonpolar side chains (e.g., alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine), β-branched side chains (e.g., threonine, valine, isoleucine), and aromatic side chains (e.g., tyrosine, phenylalanine, tryptophan, histidine). In some embodiments, one or more amino acid residues in the CDR region of the isolated antigen-binding protein described in this application may be replaced with other amino acid residues from the same side chain group. Those skilled in the art will know that some conserved sequence modifications do not cause the antigen to lose its binding ability. For example, see Brummell et al., (1993) Biochem 32:1180-8; de Wildt et al., (1997) Prot. Eng. 10:835-41; Komissarov et al., (1997) J. Biol. Chem. 272:26864-26870; Hall et al., (1992) J. Immunol. 149:1605-12; Kelley and O'Connell (1993) Biochem. 32:6862-35; Adib-Conquy et al., (1998) Int. Immunol. 10:341-6 and Beers et al., (2000) Clin. Can. Res. 6:2835-43.

[0221] The Staphylococcus aureus leukocidin antigen-binding protein described in this application can be identified, screened, or characterized by various assays known in the art.

[0222] For example, the antigen-binding activity of the antigen-binding protein of this application can be tested by known methods such as enzyme-linked immunosorbent assay (ELISA), immunoblotting (e.g., Western blotting), flow cytometry (e.g., FACS), immunohistochemistry, immunofluorescence, etc.

[0223] In this application, the isolated antigen-binding protein is capable of specifically binding to Staphylococcus aureus toxin. In this application, the Staphylococcus aureus toxin may comprise leukocidins. For example, the leukocidins may comprise HlgA, HlgB, LukD, and / or LukF. For example, the isolated antigen-binding protein is capable of specifically binding to Staphylococcus aureus HlgA, HlgB, LukD, and LukF. For example, the isolated antigen-binding protein is capable of specifically binding to Staphylococcus aureus HlgA and LukD. For example, the isolated antigen-binding protein is capable of specifically binding to Staphylococcus aureus HlgA and LukF. In this application, the isolated antigen-binding protein is capable of specifically binding to a two-component toxin, the two-component toxin being selected from a combination of F and S components, such as HlgA and HlgB, HlgA and LukD, or HlgA and LukF.

[0224] In this application, the binding of the isolated antigen-binding protein to Staphylococcus aureus leukocidin can be detected by an ELISA method. For example, the antigen-binding protein described in this application can be detected at concentrations less than or equal to about 0.1 μg / mL, less than or equal to about 0.095 μg / mL, less than or equal to about 0.09 μg / mL, less than or equal to about 0.085 μg / mL, less than or equal to about 0.08 μg / mL, less than or equal to about 0.075 μg / mL, less than or equal to about 0.06 μg / mL, less than or equal to about 0.05 μg / mL, less than or equal to about 0.04 μg / mL, less than or equal to about 0.034 μg / mL, less than or equal to about 0.03 μg / mL, less than or equal to about 0.02 μg / mL, and less than or equal to about 0.01 μg / mL. 50 The value binds to Staphylococcus aureus HlgA. In some embodiments, the antigen-binding protein described in this application binds to HlgA in EC. 50 The value is 0.076 μg / mL. In some embodiments, the antigen-binding protein described in this application binds to HClA in EC. 50 The value is 0.034 μg / mL. For example, the antigen-binding protein described in this application can be in EC values ​​less than or equal to about 0.1 μg / mL, less than or equal to about 0.09 μg / mL, less than or equal to about 0.08 μg / mL, less than or equal to about 0.07 μg / mL, less than or equal to about 0.06 μg / mL, less than or equal to about 0.055 μg / mL, less than or equal to about 0.05 μg / mL, less than or equal to about 0.045 μg / mL, less than or equal to about 0.04 μg / mL, less than or equal to about 0.035 μg / mL, and less than or equal to about 0.02 μg / mL. 50The value binds to Staphylococcus aureus HlgB. In some embodiments, the antigen-binding protein described in this application binds to HlgB in EC. 50 The value is 0.051 μg / mL. In some embodiments, the antigen-binding protein described in this application binds to HlgB in EC. 50 The value is 0.055 μg / mL. For example, the antigen-binding protein described in this application can be in EC5 values ​​less than or equal to about 0.08 μg / mL, less than or equal to about 0.07 μg / mL, less than or equal to about 0.06 μg / mL, less than or equal to about 0.05 μg / mL, less than or equal to about 0.045 μg / mL, less than or equal to about 0.04 μg / mL, less than or equal to about 0.035 μg / mL, less than or equal to about 0.034 μg / mL, less than or equal to about 0.03 μg / mL, less than or equal to about 0.02 μg / mL, and less than or equal to about 0.01 μg / mL. 50 The value binds to Staphylococcus aureus LukD. In some embodiments, the antigen-binding protein described in this application binds to LukD in EC. 50 The value is 0.038 μg / mL. In some embodiments, the antigen-binding protein described in this application binds to LukD in EC. 50 The value is 0.036 μg / mL. For example, the antigen-binding protein described in this application can be in EC5 values ​​less than or equal to about 0.08 μg / mL, less than or equal to about 0.07 μg / mL, less than or equal to about 0.06 μg / mL, less than or equal to about 0.05 μg / mL, less than or equal to about 0.045 μg / mL, less than or equal to about 0.04 μg / mL, less than or equal to about 0.035 μg / mL, less than or equal to about 0.034 μg / mL, less than or equal to about 0.03 μg / mL, less than or equal to about 0.02 μg / mL, and less than or equal to about 0.01 μg / mL. 50 The value binds to Staphylococcus aureus LukF. In some embodiments, the antigen-binding protein described in this application binds to LukF in EC. 50 The value is 0.053 μg / mL. In some embodiments, the antigen-binding protein described in this application binds to LukF in EC. 50 The value was 0.043 μg / mL.

[0225] In this application, the isolated antigen-binding protein is capable of neutralizing the activity of Staphylococcus aureus leukocidin. For example, its anti-hemolytic activity can be detected by adding rabbit erythrocytes to the Staphylococcus aureus leukocidin and antigen-binding protein and using an enzyme-linked immunosorbent assay (ELISA) reader. For example, the isolated antigen-binding protein can completely neutralize 4 μg of Staphylococcus aureus two-component toxins HlgAB, HlgA+LukD, and / or HlgA+LukF at doses of less than or equal to about 5 μg, less than or equal to about 4 μg, less than or equal to about 3 μg, less than or equal to about 2 μg, and less than or equal to about 1 μg.

[0226] In this application, the isolated antigen-binding protein is capable of preventing and / or treating diseases and / or conditions. In this application, the diseases and / or conditions may be caused or mediated by Staphylococcus aureus. In this application, the diseases and / or conditions may include complications of diseases and / or conditions caused or mediated by Staphylococcus aureus. In some embodiments, the diseases and / or conditions and their complications may include sepsis and / or bacteremia.

[0227] Polypeptide molecules, nucleic acid molecules, carriers, cells, immunoconjugates, and pharmaceutical compositions

[0228] On the other hand, this application provides polypeptide molecules that may contain the isolated antigen-binding protein described in this application.

[0229] In this application, the polypeptide molecule may comprise a fusion protein. In this application, the polypeptide molecule may be a fusion protein. In this application, the fusion protein may comprise a multispecific antibody.

[0230] On the other hand, this application provides isolated nucleic acid molecules that can encode the isolated antigen-binding protein described in this application. For example, it can be produced or synthesized by: (i) in vitro amplification, for example by polymerase chain reaction (PCR); (ii) by clonal recombination; (iii) purification, for example by enzyme digestion and gel electrophoresis fractionation; or (iv) synthesis, for example by chemical synthesis.

[0231] On the other hand, this application provides a vector that may contain the nucleic acid molecule described in this application. Furthermore, the vector may also contain other genes, such as marker genes that allow selection of the vector in appropriate host cells and under appropriate conditions. Additionally, the vector may contain expression control elements that allow the coding region to be correctly expressed in an appropriate host. Such control elements are well known to those skilled in the art and may include, for example, promoters, ribosome binding sites, enhancers, and other control elements regulating gene transcription or mRNA translation. The vector can be transformed, transduced, or transfected into host cells to express the genetic material elements it carries within the host cells. The vector may include, for example, plasmids, granules, viruses, bacteriophages, or other vectors commonly used in, for example, genetic engineering. For example, the vector is an expression vector. Furthermore, the vector may also include components that facilitate its entry into cells, such as viral particles, liposomes, or protein coats, but not only these substances.

[0232] On the other hand, this application provides a cell that may contain the nucleic acid molecules or vectors described in this application. In some embodiments, each or every host cell may contain one or more of the nucleic acid molecules or vectors described in this application. In some embodiments, each or every host cell may contain multiple (e.g., two or more) or more types (e.g., two or more) of the nucleic acid molecules or vectors described in this application. For example, the vectors described in this application may be introduced into the host cell, such as eukaryotic cells, such as cells from plants, fungi, or yeast cells. In some embodiments, the cell may be a bacterial cell (e.g., *Escherichia coli*), a yeast cell, or other eukaryotic cells, such as COS cells, Chinese hamster ovary (CHO) cells, CHO-K1 cells, LNCAP cells, HeLa cells, 293T cells, COS-1 cells, SP2 / 0 cells, NSO cells, or myeloma cells. The vectors described in this application may be introduced into the host cell by methods known in the art, such as electroporation, lipofectine transfection, lipofectamin transfection, etc.

[0233] On the other hand, this application also provides immune conjugates that may contain the isolated antigen-binding proteins described in this application.

[0234] In some embodiments, the isolated antigen-binding protein or fragment thereof described in this application can be linked to another reagent, such as a chemotherapeutic agent, toxin, immunotherapeutic agent, imaging probe, spectroscopic probe, etc. This linking can be via one or more covalent bonds, or non-covalent interactions, and may include chelation. A variety of adapters (which may be known in the art) can be used to form immunoconjugates. Furthermore, immunoconjugates can be provided in the form of fusion proteins, which may be expressed by polynucleotides encoding the immunoconjugate. The immunoconjugates may also comprise, for example, antibody-drug conjugates (ADCs). In an ADC, the antibody and the therapeutic agent can be cross-linked via a cleavable adapter, such as a peptide adapter, disulfide adapter, or hydrazone adapter.

[0235] On the other hand, this application also provides pharmaceutical compositions that may comprise the isolated antigen-binding protein described in this application, the polypeptide molecule described in this application, the immunoconjugate described in this application, the nucleic acid molecule described in this application, the carrier described in this application, and / or the cell described in this application, and optionally a pharmaceutically acceptable carrier.

[0236] In some embodiments, the pharmaceutical composition may further comprise suitable formulations of one or more (pharmaceutically effective) adjuvants, stabilizers, excipients, diluents, solubilizers, surfactants, emulsifiers, and / or preservatives. The acceptable components of the composition are preferably non-toxic to the recipient at the dosage and concentration used. The pharmaceutical compositions of the present invention include, but are not limited to, liquid, freeze-dried, and lyophilized compositions.

[0237] In some embodiments, the pharmaceutical composition may also contain more than one active compound, typically those with complementary activities that do not adversely affect each other. The type and effective amount of such a drug may depend, for example, on the amount and type of antagonist present in the formulation, and on the clinical parameters of the subject.

[0238] In some embodiments, the pharmaceutically acceptable carrier may include any and all solvents, dispersion media, coatings, isotonic agents, and absorption delay agents that are compatible with drug administration and are generally safe and non-toxic.

[0239] In some embodiments, the pharmaceutical composition may be administered parenterally, percutaneously, intracavitarily, intra-arterially, intrathecally, and / or intranasally, or directly injected into tissues. For example, the pharmaceutical composition may be administered to a patient or subject by infusion or injection. In some embodiments, the pharmaceutical composition may be administered in various ways, such as intravenously, intraperitoneally, subcutaneously, intramuscularly, locally, or intradermally. In some embodiments, the pharmaceutical composition may be administered continuously. This continuous (or uninterrupted) administration may be achieved using a small pump system worn by the patient to measure the amount of therapeutic agent flowing into the patient, as described in WO2015 / 036583.

[0240] Preparation method

[0241] On the other hand, this application provides a method for preparing the isolated antigen-binding protein. The method may include culturing the host cells described in this application under conditions that allow the antigen-binding protein to be expressed. For example, this can be achieved by using appropriate culture media, appropriate temperatures, and culture times, methods known to those skilled in the art.

[0242] Any method suitable for generating monoclonal antibodies can be used to generate the antigen-binding protein of this application. For example, animals can be immunized with either conjugated or naturally occurring Staphylococcus aureus leukocidin or fragments thereof. Suitable immunization methods can be used, including adjuvants, immunostimulants, repeated booster immunizations, or one or more pathways. In some embodiments, the isolated antigen-binding protein against Staphylococcus aureus leukocidin can be screened and enriched by extracting peripheral blood lymphocytes from immunized alpaca, cloning the cellular nucleic acid fragment into a vector, and then using a phage surface display system.

[0243] Any suitable form of Staphylococcus aureus leukocidin can serve as an immunogen (antigen) for generating non-human antibodies specific to Staphylococcus aureus leukocidin and screening for the biological activity of said antibodies. For example, the stimulating immunogen can be full-length Staphylococcus aureus leukocidin, including a natural homodimer or a peptide containing one or more epitopes. The immunogen can be used alone or in combination with one or more immunogenicity enhancers known in the art.

[0244] Methods and uses

[0245] On the other hand, this application provides the use of the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate and / or the pharmaceutical composition in the preparation of a medicament for the prevention and / or treatment of diseases and / or conditions.

[0246] On the other hand, this application also provides methods for preventing and / or treating diseases and / or conditions, which may include administering to a subject in need the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate, and / or the pharmaceutical composition described in this application.

[0247] In this application, the application can be performed in various ways, such as intravenous, intratumoral, intraperitoneal, subcutaneous, intramuscular, local, or intradermal application.

[0248] On the other hand, the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate, and / or the pharmaceutical composition described in this application can be used to prevent and / or treat diseases and / or conditions.

[0249] In this application, the disease and / or condition may be caused or mediated by Staphylococcus aureus.

[0250] In this application, the disease and / or condition may be a complication of a disease and / or condition caused or mediated by Staphylococcus aureus.

[0251] In this application, the disease and / or ailment may include bacterial infection.

[0252] In this application, the disease and / or ailment may include bacteremia and / or sepsis.

[0253] In this application, the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate, and / or the pharmaceutical composition may be used alone or in combination with other drugs to prevent and / or treat diseases and / or conditions. In some embodiments, the other drugs may be any currently known antibacterial agents. The diseases and / or conditions may be caused by Staphylococcus aureus infection or related diseases caused by other infections.

[0254] On the other hand, this application also provides a method for detecting Staphylococcus aureus leukocidin in a sample, the method comprising administering the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate and / or the pharmaceutical composition.

[0255] In this application, the method for detecting Staphylococcus aureus leukocidin in a sample can be an in vitro method. For example, the isolated antigen-binding protein described in this application is brought into contact with an isolated sample to detect the presence and / or content of Staphylococcus aureus leukocidin in the sample. In some cases, the method for detecting Staphylococcus aureus leukocidin in a sample is for non-therapeutic purposes. In some cases, the method for detecting Staphylococcus aureus leukocidin in a sample is not a diagnostic method.

[0256] On the other hand, this application also provides a reagent or kit for detecting Staphylococcus aureus leukocidin in a sample, which comprises the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate and / or the pharmaceutical composition.

[0257] On the other hand, this application also provides the use of the isolated antigen-binding protein, the polypeptide molecule, the nucleic acid molecule, the carrier, the cell, the immunoconjugate and / or the pharmaceutical composition in the preparation of a kit for detecting the presence and / or content of Staphylococcus aureus leukocidin in a sample.

[0258] This application also includes the following specific implementation schemes:

[0259] 1. An isolated antigen-binding protein comprising at least one CDR in the variable region VH of an antibody heavy chain, said VH comprising the amino acid sequence shown in SEQ ID NO:53.

[0260] 2. The isolated antigen-binding protein according to embodiment 1, comprising HCDR3, wherein HCDR3 comprises the amino acid sequence shown in SEQ ID NO:1.

[0261] 3. The isolated antigen-binding protein according to any one of embodiments 1-2, comprising HCDR2, wherein the HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2.

[0262] 4. The isolated antigen-binding protein according to any one of embodiments 1-3, comprising HCDR1, wherein HCDR1 comprises the amino acid sequence shown in SEQ ID NO:3.

[0263] 5. The isolated antigen-binding protein according to any one of embodiments 1-4, comprising HCDR1, HCDR2 and HCDR3, wherein HCDR1 comprises the amino acid sequence shown in SEQ ID NO:3, HCDR2 comprises the amino acid sequence shown in SEQ ID NO:2 and HCDR3 comprises the amino acid sequence shown in SEQ ID NO:1.

[0264] 6. The isolated antigen-binding protein according to any one of embodiments 4-5, comprising H-FR1, wherein the C-terminus of H-FR1 is directly or indirectly linked to the N-terminus of HCDR1, and wherein H-FR1 comprises the amino acid sequence shown in SEQ ID NO:45.

[0265] 7. The isolated antigen-binding protein according to embodiment 6, wherein the H-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:10, SEQ ID NO:14 and SEQ ID NO:18.

[0266] 8. The isolated antigen-binding protein according to any one of embodiments 4-7, comprising H-FR2 located between HCDR1 and HCDR2, and wherein H-FR2 comprises the amino acid sequence shown in SEQ ID NO:46.

[0267] 9. The isolated antigen-binding protein according to embodiment 8, wherein the H-FR2 comprises the amino acid sequence shown in any one of SEQ ID NO:11, SEQ ID NO:15 and SEQ ID NO:19.

[0268] 10. The isolated antigen-binding protein according to any one of embodiments 3-9, comprising H-FR3 located between HCDR2 and HCDR3, and wherein the H-FR3 comprises the amino acid sequence shown in SEQ ID NO:47.

[0269] 11. According to the amino acid sequence shown in Embodiment 10, the H-FR3 comprises the amino acid sequence shown in any one of SEQ ID NO:12, SEQ ID NO:16 and SEQ ID NO:20.

[0270] 12. The isolated antigen-binding protein according to any one of embodiments 2-11, comprising H-FR4, wherein the N-terminus of H-FR4 is linked to the C-terminus of HCDR3, and wherein H-FR4 comprises the amino acid sequence shown in SEQ ID NO:48.

[0271] 13. The isolated antigen-binding protein according to embodiment 12, wherein the H-FR4 comprises the amino acid sequence shown in SEQ ID NO:13 or SEQ ID NO:17.

[0272] 14. The isolated antigen-binding protein according to any one of embodiments 1-13, comprising H-FR1, H-FR2, H-FR3 and H-FR4, wherein H-FR1 comprises the amino acid sequence shown in SEQ ID NO:45, H-FR2 comprises the amino acid sequence shown in SEQ ID NO:46, H-FR3 comprises the amino acid sequence shown in SEQ ID NO:47, and H-FR4 comprises the amino acid sequence shown in SEQ ID NO:48.

[0273] 15. The isolated antigen-binding protein according to any one of embodiments 1-14, comprising H-FR1, H-FR2, H-FR3 and H-FR4, wherein H-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:10, SEQ ID NO:14 and SEQ ID NO:18, H-FR2 comprises the amino acid sequence shown in any one of SEQ ID NO:11, SEQ ID NO:15 and SEQ ID NO:19, H-FR3 comprises the amino acid sequence shown in any one of SEQ ID NO:12, SEQ ID NO:16 and SEQ ID NO:20, and H-FR4 comprises the amino acid sequence shown in SEQ ID NO:13 or SEQ ID NO:17.

[0274] 16. The isolated antigen-binding protein according to any one of embodiments 1-15, comprising H-FR1, H-FR2, H-FR3 and H-FR4 selected from any one of the following groups:

[0275] 1) H-FR1 contains the amino acid sequence shown in SEQ ID NO:10, H-FR2 contains the amino acid sequence shown in SEQ ID NO:11, H-FR3 contains the amino acid sequence shown in SEQ ID NO:12, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:13;

[0276] 2) H-FR1 contains the amino acid sequence shown in SEQ ID NO:14, H-FR2 contains the amino acid sequence shown in SEQ ID NO:15, H-FR3 contains the amino acid sequence shown in SEQ ID NO:16, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17;

[0277] 3) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:15, H-FR3 contains the amino acid sequence shown in SEQ ID NO:16, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17;

[0278] 4) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:19, H-FR3 contains the amino acid sequence shown in SEQ ID NO:16, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17;

[0279] 5) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:15, H-FR3 contains the amino acid sequence shown in SEQ ID NO:20, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17; and

[0280] 6) H-FR1 contains the amino acid sequence shown in SEQ ID NO:18, H-FR2 contains the amino acid sequence shown in SEQ ID NO:19, H-FR3 contains the amino acid sequence shown in SEQ ID NO:20, and H-FR4 contains the amino acid sequence shown in SEQ ID NO:17.

[0281] 17. The isolated antigen-binding protein according to any one of embodiments 1-16, comprising VH, wherein the VH comprises the amino acid sequence shown in SEQ ID NO:53.

[0282] 18. The isolated antigen-binding protein according to embodiment 17, wherein the VH comprises the amino acid sequence shown in any one of SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:34 and SEQ ID NO:35.

[0283] 19. The isolated antigen-binding protein according to any one of embodiments 1-18, comprising the constant region of the antibody heavy chain.

[0284] 20. The isolated antigen-binding protein according to embodiment 19, wherein the heavy chain constant region is derived from the human IgG constant region.

[0285] 21. The isolated antigen-binding protein according to any one of embodiments 19-20, wherein the heavy chain constant region is derived from the human IgG1 heavy chain constant region.

[0286] 22. The isolated antigen-binding protein according to any one of embodiments 19-21, wherein the heavy chain constant region comprises the amino acid sequence shown in SEQ ID NO:42.

[0287] 23. The isolated antigen-binding protein according to any one of embodiments 1-21, comprising an antibody heavy chain, said heavy chain comprising the amino acid sequence shown in any one of SEQ ID NO:55, SEQ ID NO:57, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:60 and SEQ ID NO:61.

[0288] 24. The isolated antigen-binding protein according to any one of embodiments 1-23, comprising at least one CDR in the variable region VL of the antibody light chain, said VL comprising the amino acid sequence shown in SEQ ID NO:54.

[0289] 25. The isolated antigen-binding protein according to any one of embodiments 1-24, comprising LCDR3, wherein LCDR3 comprises the amino acid sequence shown in SEQ ID NO:4.

[0290] 26. The isolated antigen-binding protein according to any one of embodiments 1-25, comprising LCDR2, wherein LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5.

[0291] 27. The isolated antigen-binding protein according to any one of embodiments 1-26, comprising LCDR1, wherein LCDR1 comprises the amino acid sequence shown in SEQ ID NO:44.

[0292] 28. The isolated antigen-binding protein according to embodiment 27, wherein the LCDR1 comprises the amino acid sequence shown in any one of SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8 and SEQ ID NO:9.

[0293] 29. The isolated antigen-binding protein according to any one of embodiments 1-28, comprising LCDR1, LCDR2 and LCDR3, wherein LCDR1 comprises the amino acid sequence shown in SEQ ID NO:44, LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5 and LCDR3 comprises the amino acid sequence shown in SEQ ID NO:4.

[0294] 30. The isolated antigen-binding protein according to any one of embodiments 1-29, comprising LCDR1, LCDR2 and LCDR3, wherein LCDR1 comprises the amino acid sequence shown in any one of SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8 and SEQ ID NO:9, LCDR2 comprises the amino acid sequence shown in SEQ ID NO:5, and LCDR3 comprises the amino acid sequence shown in SEQ ID NO:4.

[0295] 31. The isolated antigen-binding protein according to any one of embodiments 27-30, comprising L-FR1, wherein the C-terminus of L-FR1 is directly or indirectly linked to the N-terminus of LCDR1, and wherein L-FR1 comprises the amino acid sequence shown in SEQ ID NO:49.

[0296] 32. The isolated antigen-binding protein according to embodiment 31, wherein the L-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:21, SEQ ID NO:25 and SEQ ID NO:29.

[0297] 33. The isolated antigen-binding protein according to any one of embodiments 27-32, comprising L-FR2 located between LCDR1 and LCDR2, and wherein L-FR2 comprises the amino acid sequence shown in SEQ ID NO:50.

[0298] 34. The isolated antigen-binding protein according to embodiment 33, wherein the L-FR2 comprises the amino acid sequence shown in SEQ ID NO:22 or SEQ ID NO:26.

[0299] 35. The isolated antigen-binding protein according to any one of embodiments 26-34, comprising L-FR3 located between LCDR2 and LCDR3, and wherein the L-FR3 comprises the amino acid sequence shown in SEQ ID NO:51.

[0300] 36. The isolated antigen-binding protein according to embodiment 35, wherein the L-FR3 comprises the amino acid sequence shown in SEQ ID NO:23 or SEQ ID NO:27.

[0301] 37. The isolated antigen-binding protein according to any one of embodiments 1-36, comprising L-FR4, wherein the N-terminus of L-FR4 is directly or indirectly linked to the C-terminus of LCDR3, and wherein L-FR4 comprises the amino acid sequence shown in SEQ ID NO:52.

[0302] 38. The isolated antigen-binding protein according to embodiment 37, wherein the L-FR4 comprises the amino acid sequence shown in SEQ ID NO:24 or SEQ ID NO:28.

[0303] 39. The isolated antigen-binding protein according to any one of embodiments 1-38, comprising L-FR1, L-FR2, L-FR3 and L-FR4, wherein L-FR1 comprises the amino acid sequence shown in SEQ ID NO:49, L-FR2 comprises the amino acid sequence shown in SEQ ID NO:50, L-FR3 comprises the amino acid sequence shown in SEQ ID NO:51, and L-FR4 comprises the amino acid sequence shown in SEQ ID NO:52.

[0304] 40. The isolated antigen-binding protein according to any one of embodiments 1-39, comprising L-FR1, L-FR2, L-FR3 and L-FR4, wherein L-FR1 comprises the amino acid sequence shown in any one of SEQ ID NO:21, SEQ ID NO:25 and SEQ ID NO:29, L-FR2 comprises the amino acid sequence shown in SEQ ID NO:22 or SEQ ID NO:26, L-FR3 comprises the amino acid sequence shown in SEQ ID NO:23 or SEQ ID NO:27, and L-FR4 comprises the amino acid sequence shown in SEQ ID NO:24 or SEQ ID NO:28.

[0305] 41. The isolated antigen-binding protein according to any one of embodiments 1-40, comprising L-FR1, L-FR2, L-FR3 and L-FR4 selected from any one of the following groups:

[0306] 1) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:21, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:22, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:23, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:24;

[0307] 2) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:25, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:26, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:27, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:28; and

[0308] 3) The L-FR1 contains the amino acid sequence shown in SEQ ID NO:29, the L-FR2 contains the amino acid sequence shown in SEQ ID NO:26, the L-FR3 contains the amino acid sequence shown in SEQ ID NO:27, and the L-FR4 contains the amino acid sequence shown in SEQ ID NO:28.

[0309] 42. The isolated antigen-binding protein according to any one of embodiments 1-41, comprising VL, wherein the VL comprises the amino acid sequence shown in SEQ ID NO:54.

[0310] 43. The isolated antigen-binding protein according to embodiment 42, wherein the VL comprises the amino acid sequence shown in any one of SEQ ID NO:36, SEQ ID NO:37, SEQ ID NO:38, SEQ ID NO:39, SEQ ID NO:40 and SEQ ID NO:41.

[0311] 44. The isolated antigen-binding protein according to any one of embodiments 1-43, comprising the constant region of the antibody light chain.

[0312] 45. The isolated antigen-binding protein according to embodiment 44, wherein the light chain constant region is derived from the human Igκ constant region.

[0313] 46. ​​The isolated antigen-binding protein according to any one of embodiments 44-45, wherein the light chain constant region comprises the amino acid sequence shown in SEQ ID NO:43.

[0314] 47. The isolated antigen-binding protein according to any one of embodiments 1-45, comprising an antibody light chain, said light chain comprising the amino acid sequence shown in any one of SEQ ID NO:56, SEQ ID NO:62, SEQ ID NO:63, SEQ ID NO:64, SEQ ID NO:65 and SEQ ID NO:66.

[0315] 48. The isolated antigen-binding protein according to any one of embodiments 1-47, comprising VH and VL selected from any one of the following groups:

[0316] 1) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0317] 2) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0318] 3) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0319] 4) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0320] 5) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0321] 6) The VH contains the amino acid sequence shown in SEQ ID NO:30, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0322] 7) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0323] 8) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0324] 9) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0325] 10) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0326] 11) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0327] 12) The VH contains the amino acid sequence shown in SEQ ID NO:31, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0328] 13) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0329] 14) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0330] 15) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0331] 16) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0332] 17) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0333] 18) The VH contains the amino acid sequence shown in SEQ ID NO:32, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0334] 19) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0335] 20) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0336] 21) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0337] 22) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0338] 23) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0339] 24) The VH contains the amino acid sequence shown in SEQ ID NO:33, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0340] 25) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0341] 26) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0342] 27) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0343] 28) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0344] 29) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:40;

[0345] 30) The VH contains the amino acid sequence shown in SEQ ID NO:34, and the VL contains the amino acid sequence shown in SEQ ID NO:41;

[0346] 31) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:36;

[0347] 32) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:37;

[0348] 33) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:38;

[0349] 34) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:39;

[0350] 35) The VH comprises the amino acid sequence shown in SEQ ID NO:35, and the VL comprises the amino acid sequence shown in SEQ ID NO:40; and

[0351] 36) The VH contains the amino acid sequence shown in SEQ ID NO:35, and the VL contains the amino acid sequence shown in SEQ ID NO:41.

[0352] 49. An isolated antigen-binding protein according to any one of embodiments 1-48, comprising an antibody or an antigen-binding fragment thereof.

[0353] 50. The isolated antigen-binding protein according to embodiment 49, wherein the antigen-binding fragment includes Fab, Fab', Fv fragment, F(ab')2, F(ab)2, scFv, di-scFv and / or dAb.

[0354] 51. The isolated antigen-binding protein according to any one of embodiments 49-50, wherein the antibody is selected from one or more of the group consisting of monoclonal antibodies, chimeric antibodies, humanized antibodies, and fully human antibodies.

[0355] 52. The isolated antigen-binding protein according to any one of embodiments 1-51, which is capable of specifically binding to Staphylococcus aureus toxin.

[0356] 53. The isolated antigen-binding protein according to embodiment 52, wherein the Staphylococcus aureus toxin comprises leukocidin.

[0357] 54. The isolated antigen-binding protein according to any one of embodiments 52-53, wherein the Staphylococcus aureus toxin is selected from one or more of the group consisting of HlgA, HlgB, LukD and LukF.

[0358] 55. The isolated antigen-binding protein according to any one of embodiments 1-54, which is capable of neutralizing the activity of one or more Staphylococcus aureus virulence factors selected from the group consisting of HlgA, HlgB, LukD and LukF.

[0359] 56. The isolated antigen-binding protein according to any one of embodiments 1-55, which is capable of treating diseases and / or symptoms caused by Staphylococcus aureus.

[0360] 57. The isolated antigen-binding protein according to embodiment 56, wherein the disease and / or condition includes complications of disease and / or condition caused by Staphylococcus aureus.

[0361] 58. A polypeptide molecule comprising the isolated antigen-binding protein of any one of embodiments 1-57.

[0362] 59. The polypeptide molecule according to embodiment 58, comprising a fusion protein.

[0363] 60. An immunoconjugate comprising the isolated antigen-binding protein of any one of embodiments 1-57.

[0364] 61. A nucleic acid molecule encoding an isolated antigen-binding protein as described in any one of embodiments 1-57 or a polypeptide molecule as described in any one of embodiments 58-59.

[0365] 62. A vector comprising the nucleic acid molecule described in embodiment 61.

[0366] 63. A cell comprising the nucleic acid molecule described in embodiment 61 or the vector described in embodiment 62.

[0367] 64. A pharmaceutical composition comprising the isolated antigen-binding protein of any one of embodiments 1-57, the polypeptide molecule of any one of embodiments 58-59, the immunoconjugate of embodiment 60, the nucleic acid molecule of embodiment 61, the carrier of embodiment 62 and / or the cell of embodiment 63, and optionally a pharmaceutically acceptable carrier.

[0368] 65. A method for preparing the isolated antigen-binding protein according to any one of embodiments 1-57, the method comprising culturing the cells described in embodiment 63 under conditions that cause the antigen-binding protein to be expressed.

[0369] 66. Use of the isolated antigen-binding protein of any one of embodiments 1-57, the polypeptide molecule of any one of embodiments 58-59, the immunoconjugate of embodiment 60, the nucleic acid molecule of embodiment 61, the carrier of embodiment 62, the cell of embodiment 63, and / or the pharmaceutical composition of embodiment 64 in the preparation of a medicament for the prevention and / or treatment of diseases and / or conditions.

[0370] 67. The use according to embodiment 65, wherein the disease and / or condition and its complications are caused or mediated by Staphylococcus aureus.

[0371] 68. The use according to any one of embodiments 66-67, wherein the disease and / or condition includes complications of diseases and / or conditions caused or mediated by Staphylococcus aureus.

[0372] 69. The use according to any one of embodiments 66-68, wherein the disease and / or symptom includes sepsis and / or bacteremia.

[0373] 70. The isolated antigen-binding protein of any one of embodiments 1-57, the polypeptide molecule of any one of embodiments 58-59, the immunoconjugate of embodiment 60, the nucleic acid molecule of embodiment 61, the carrier of embodiment 62, the cell of embodiment 63, and / or the pharmaceutical composition of embodiment 64, used alone or in combination with other drugs.

[0374] 71. A method for detecting Staphylococcus aureus leukocidin in a sample, the method comprising administering the isolated antigen-binding protein of any one of embodiments 1-57, the polypeptide molecule of any one of embodiments 58-59, the immunoconjugate of embodiment 60, the nucleic acid molecule of embodiment 61, the carrier of embodiment 62, the cell of embodiment 63, and / or the pharmaceutical composition of embodiment 64.

[0375] 72. A kit for detecting Staphylococcus aureus leukocidin in a sample, said kit comprising the isolated antigen-binding protein of any one of embodiments 1-57, the polypeptide molecule of any one of embodiments 58-59, the immunoconjugate of embodiment 60, the nucleic acid molecule of embodiment 61, the carrier of embodiment 62, the cell of embodiment 63, and / or the pharmaceutical composition of embodiment 64.

[0376] 73. Use of the isolated antigen-binding protein of any one of embodiments 1-57, the polypeptide molecule of any one of embodiments 58-59, the immunoconjugate of embodiment 60, the nucleic acid molecule of embodiment 61, the carrier of embodiment 62, the cell of embodiment 63, and / or the pharmaceutical composition of embodiment 64 in the preparation of a kit for detecting the presence and / or content of Staphylococcus aureus leukocidin in a sample.

[0377] The embodiments described below are not intended to be limited by any theory, but are merely for illustrating the antigen-binding protein, preparation method and use of this application, and are not intended to limit the scope of the invention.

[0378] Example

[0379] Example 1: Recombinant Expression of Staphylococcus aureus Leukocidin

[0380] Recombinant Staphylococcus aureus plasmids HlgA (Uniport KB-P0A074, 30-309, C-terminal His tag), HlgB (Uniport KB-P0A077, 26-325, C-terminal His tag), LukD (Uniport KB-O54082, 27-327, C-terminal His tag), and LukF (UniProt KB-P31715, 26-323, C-terminal His tag) were constructed. After transforming the recombinant plasmids into BL21(DE3)pLysS competent cells, protein expression was performed using an Escherichia coli expression system. The supernatant from ultrasonic lysis of the bacterial culture was purified by Ni-NTA affinity chromatography to obtain HlgA, HlgB, LukD, and LukF proteins.

[0381] Example 2: Immunization of Balb / C mice with a mixture of HlgA and LukF antigens

[0382] Five Balb / C mice were subcutaneously immunized at multiple sites using the prepared HlgA and LukF proteins.

[0383] The immunization process is as follows:

[0384]

[0385] Example 3: Fusion of spleen cells and myeloma cells in immunized mice and screening of antigen-binding proteins with cross-affinity.

[0386] Myeloma cells P3X63Ag8.653 were fused with spleen cells from immunized HLG and LukF mice using PEG3350. The fused hybridoma cells were seeded into 96-well plates and cultured. After 24 and 48 hours, media containing HAT and HT were added, respectively, to screen for hybridoma cells. After 12 days of culture, 100 μL of cell culture supernatant was collected. Hybridoma clones capable of producing anti-HLG and LukF antigen-binding proteins were screened using ELISA. Positive hybridoma cell clusters were added to 96-well plates containing mouse spleen cells using a limiting dilution method, and single-clone cells were labeled. Further screening using ELISA revealed single-clone cells capable of secreting anti-HLG and LukF antigen-binding proteins.

[0387] Example 4: Obtaining the sequence of a murine antigen-binding protein with cross-affinity

[0388] 1×10⁻⁶ RNA was extracted using the RNAfast200 kit (Shanghai Feijie Biotechnology Co., Ltd.) 6 Total RNA from monoclonal cells secreting anti-HLG and LukF antigen-binding proteins was obtained. The total RNA was reverse transcribed into cDNA using a reverse transcription kit (Takara). The cDNA fragment was amplified using primers (Anke Krebber. 1997), and the PCR product was purified using a DNA product purification kit (Beijing Tiangen Biotech Co., Ltd.). The PCR product was then ligated into a T-vector using a ligation kit (Beijing Qingke Biotechnology Co., Ltd.) and transformed into competent cells. Sequencing was performed by Shanghai Sangon Biotech Co., Ltd., ultimately obtaining the heavy chain and light chain variable regions of the murine antigen-binding protein 17C7.

[0389] 17C7 heavy chain variable region (17C7Hv) SEQ ID NO:30

[0390] EVQLQQSGAELVRSGASVKLSCTASGFNIK DYYMHWVKQRPEQGLEWIG WIDPENDI GEYAPKFQG KATMTADTSSNTAYLQLSSLTSEDTAVYFCNA LWGGVLDY WGQGTSVTVSA

[0391] 17C7 light chain variable region (17C7Lv) SEQ ID NO:36

[0392] DVVMTQTPLSLPVSLGDQASISC RSSQSLVNSNGNTYLH WYLQKPGQSPNLLIY RVSN RFS GVPDRFSGSGSGTDFTLKISRVEAEDLGVYFC SQSTHVPYT FGGGTKLEIKR

[0393] The underlined portion represents the CDR region defined using the Kabat method.

[0394] Example 5: ELISA method for detecting the binding of antigen-binding proteins to various Staphylococcus aureus leukocidins

[0395] Dilute leukocytocin to 1 μg / mL with PBS buffer, and coat each well with 100 μL of the solution in a 96-well plate (Thermo) and incubate overnight at 4°C. The next day, remove the 96-well plate, wash with PBST (containing 0.5% PBS), soaking for 1 min each time and then thoroughly dry. Add 200 μL of PBST containing 5% BSA to each sample well and block at 37°C for 1 h; then wash with PBST and dry the wells. Add 100 μL of the test sample to each 96-well plate and incubate overnight at 4°C. After removing the 96-well plate, wash with PBST and add 100 μL of secondary antibody (HRP antibody, Beijing Dingguo Biotechnology Co., Ltd.) to each well, then incubate at 37°C for 1 h. Wash 5 times with PBST, add 100 μL of Substrate Solution (Invitrogen) to each well, and incubate at 37 °C for 10 min. After stopping the reaction, add 50 μL of 1 mol sulfuric acid to each well and then measure the absorbance at 450 nm using a microplate reader (Multiskcin FC, Thermo).

[0396] Test results as follows Figure 1 As shown, the results indicate that the antigen-binding protein described in this application can bind to leukocidins HlgA, HlgB, LukD, and LukF. EC20 binding to HlgA... 50 The value was 0.076 μg / mL, and the EC binding to HlgB was... 50 The value was 0.051 μg / mL, and the EC bound to LukD was... 50The value was 0.038 μg / mL, and the EC bound to LukF was... 50 The value was 0.053 μg / mL.

[0397] Example 6: In vitro neutralization of multiple Staphylococcus aureus leukocidin activity assay

[0398] Leukocytocin was serially diluted to 5 μg / mL to prepare a stock solution. 2 μg of leukocytocin was combined with leukocytocin antigen-binding protein (0.25–4 μg), PBS, and 5% rabbit erythrocytes and added to a 96-well plate, then mixed thoroughly. The entire mixture was incubated at 37°C for 1 h, followed by centrifugation to obtain the supernatant. 100 μl of the supernatant was transferred to a new 96-well plate, and the OD405 absorbance was measured using a microplate reader.

[0399] Test results as follows Figure 2 As shown, the results indicate that 4 μg of 17C7 antigen-binding protein completely neutralizes 4 μg of the combined two-component toxins HlgAB(A), HlgA+LukD(B), and HlgA+LukF(C).

[0400] Example 7: Protective effect of murine antigen-binding protein 17C7 against Staphylococcus aureus infection in mice.

[0401] Methicillin-resistant Staphylococcus aureus (MRSA) seed bank (USA300) was retrieved, activated for two generations on TSA solid agar plates, and then inoculated into TSB liquid agar for overnight culture. The bacterial cells were collected by centrifugation and resuspended in PBS for later use. C57BL / 6J mice were randomly divided into a model group and an antigen-binding protein drug treatment group based on body weight. Mice in each group were infected with USA300 6 × 10⁶ cells via tail vein. 7 CFU / mouse, and administered appropriate drug treatment 2 hours after infection; antigen-binding protein and control antibody AR-301 were administered at a dose of 15 mg / kg (tail vein injection). AR-301 is a fully human antibody drug (Salvecin, tosatoxumab) developed by Aridis Pharmaceuticals that targets Staphylococcus aureus α-toxin. The survival status of mice in each group was observed and recorded.

[0402] Based on the experimental results of bacteremia induced by Staphylococcus aureus in mice ( Figure 3 The results showed that antigen-binding protein 17C7 had a significant protective effect against Staphylococcus aureus-induced bacteremia in mice, and was superior to the control antibody AR-301.

[0403] Example 8 Humanization of mouse antigen-binding protein 17C7

[0404] By comparing with the NCBI database (https: / / www.ncbi.nlm.nih.gov / igblast / ), the germline sequence closest to the 17C7 antigen-binding protein was selected as a template to humanize 17C7, and the post-translational modification sites of the antigen-binding protein were optimized to obtain the humanized sequence as follows.

[0405] Humanized antigen-binding protein 17C7 heavy chain variable region:

[0406] 17C7H3v SEQ ID NO:31

[0407] QVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWIG WIDPENDIGEYAPKFQG RATMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSS

[0408] 17C7H4v SEQ ID NO:32

[0409] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWIG WIDPENDIGEYAPKFQG RATMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSS

[0410] 17C7H12v SEQ ID NO:33

[0411] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWMG WIDPENDIGEYAPKFQG RATMTADTSISTAYMELSRLRSDDTAVYYCNALWGGVLDYWGQGTLVTVSS

[0412] 17C7H13v SEQ ID NO:34

[0413] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWIG WIDPENDIGEYAPKFQG RVTMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSS

[0414] 17C7H17v SEQ ID NO:35

[0415] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWMG WIDPENDIGEYAPKFQG RVTMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSS

[0416] Humanized antigen-binding protein 17C7 light chain variable region:

[0417] 17C7L4v SEQ ID NO:37

[0418] DVVMTQTPLSSPVTLGQPASISC RSSQSLVASNGNTYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKR

[0419] 17C7L5v SEQ ID NO:38

[0420] DVVMTQTPLSSPVTLGQPASISC RSSQSLVASNRNTYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKR

[0421] 17C7L6v SEQ ID NO:39

[0422] DVVMTQTPLSSPVTLGQPASISC RSSQSLVASNRATYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKR

[0423] 17C7L7v SEQ ID NO:40

[0424] DIVMTQTPLSSPVTLGQPASISC RSSQSLVASNRNTYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKR

[0425] 17C7L8v SEQ ID NO:41

[0426] DIVMTQTPLSSPVTLGQPASISC RSSQSLVASNRATYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKR

[0427] The underlined portion represents the CDR region defined using the Kabat method.

[0428] Example 9 Recombinant Expression and Purification of 17C7 Humanized Antigen-Binding Protein

[0429] The C-terminus of the variable region of the 17C7 heavy chain was linked to the N-terminus of the constant region of human immunoglobulin γ1 (IgG1) and constructed into the pcDNA3.4 vector to obtain the 17C7 chimeric antibody heavy chain expression vector pcDNA3.4-17C7H. The C-terminus of the variable region of the 17C7 light chain was linked to the N-terminus of the kappa constant region of the light chain and constructed into the pcDNA3.4 vector to obtain the 17C7 chimeric antibody light chain expression vector pcDNA3.4-17C7L.

[0430] SEQ ID NO:55 of the 17C7 chimeric antibody heavy chain (17C7H) sequence

[0431] EVQLQQSGAELVRSGASVKLSCTASGFNIK DYYMH WVKQRPEQGLEWIG WIDPENDIGEYAPKFQG KATMTADTSSNTAYLQLSSLTSEDTAVYFCNA LWGGVLDYWGQGTSVTVSAASTKGPSVFPLAPSSKSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFN WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK

[0432] SEQ ID NO:56 of the 17C7 chimeric antibody light chain (17C7L) sequence

[0433] DVVMTQTPLSLPVSLGDQASISC RSSQSLVNSNGNTYLH WYLQKPGQSPNLLIY RVSNRFS GVPDRFSGSGSGTDFTLKISRVEAEDLGVYFC SQSTHVPYT FGGGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0434] The C-terminus of the variable regions 17C7H3v, 17C7H4v, 17C7H12v, 17C7H13v, and 17C7H17v of the humanized heavy chain were linked to the N-terminus of the constant region of human immunoglobulin γ1 (IgG1) and constructed into the pcDNA3.4 vector to obtain the humanized heavy chain expression vectors pcDNA3.4-17C7H3, pcDNA3.4-17C7H4, pcDNA3.4-17C7H12, pcDNA3.4-17C7H13, and pcDNA3.4-17C7H17.

[0435] Humanized antigen-binding protein 17C7 heavy chain sequence:

[0436] 17C7H3 SEQ ID NO:57

[0437] QVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWIG WIDPENDIGEYAPKFQG RATMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0438] 17C7H4 SEQ ID NO:58

[0439] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWIG WIDPENDIGEYAPKFQG RATMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0440] 17C7H12 SEQ ID NO:59

[0441] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWMG WIDPENDIGEYAPKFQG RATMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDY WGQGTLVTVSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0442] 17C7H13 SEQ ID NO:60

[0443] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWIG WIDPENDIGEYAPKFQG RVTMTADTSISTAYMELSRLRSDDTAVYYCNA LWGGVLDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0444] 17C7H17 SEQ ID NO:61

[0445] EVQLVQSGAEVKKPGASVKVSCKASGFNIK DYYMH WVRQAPGQGLEWMG WIDPENDIGEYAPKFQG RVTMTADTSISTAYMELSRLRSDDTAVYCNA LWGGVLDY WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLYITREPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPG

[0446] The C-terminus of the variable regions 17C7L4v, 17C7L5v, 17C7L6v, 17C7L7v, and 17C7L8v of the humanized light chain were linked to the N-terminus of the kappa constant region of the light chain and constructed into the pcDNA3.4 vector to obtain the humanized light chain expression vectors pcDNA3.4-17C7L4, pcDNA3.4-17C7L5, pcDNA3.4-17C7L6, pcDNA3.4-17C7L7, and pcDNA3.4-17C7L8.

[0447] Humanized antigen-binding protein 17C7 light chain sequence:

[0448] 17C7L4 SEQ ID NO:62

[0449] DVVMTQTPLSSPVTLGQPASISC RSSQSLVASNGNTYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0450] 17C7L5 SEQ ID NO:63

[0451] DVVMTQTPLSSPVTLGQPASISC RSSQSLVASNRNTYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0452] 17C7L6 SEQ ID NO:64

[0453] DVVMTQTPLSSPVTLGQPASISC RSSQSLVASNRATYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYTFGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0454] 17C7L7 SEQ ID NO:65

[0455] DIVMTQTPLSSPVTLGQPASISC RSSQSLVASNRNTYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0456] 17C7L8 SEQ ID NO:66

[0457] DIVMTQTPLSSPVTLGQPASISC RSSQSLVASNRATYLH WYQQRPGQPPRLLIY RVSNRFS GVPDRFSGSGAGTDFTLKISRVEAEDVGVYYC SQSTHVPYT FGQGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC

[0458] Heavy chain expression vectors pcDNA3.4-17C7H, pcDNA3.4-17C7H3, pcDNA3.4-17C7H4, pcDNA3.4-17C7H12, pcDNA3.4-17C7H13, or pcDNA3.4-17C7H17 were co-transfected with light chain expression vectors pcDNA3.4-17C7L, pcDNA3.4-17C7L4, pcDNA3.4-17C7L5, pcDNA3.4-17C7L6, pcDNA3.4-17C7L7, or pcDNA3.4-17C7L8 into expi293F cells for expression. The cells were then purified using Protein A magnetic beads to obtain the purified chimeric antibody 17C7HL and various humanized antibodies.For example, co-transfecting cells with the heavy chain expression vector pcDNA3.4-17C7H and the light chain expression vector pcDNA3.4-17C7L5 yields the humanized antibody 17C7HL5; co-transfecting cells with the heavy chain expression vector pcDNA3.4-17C7H and the light chain expression vector pcDNA3.4-17C7L6 yields the humanized antibody 17C7HL6; co-transfecting cells with the heavy chain expression vector pcDNA3.4-17C7H and the light chain expression vector pcDNA3.4-17C7L7 yields the humanized antibody 17C7HL7; and co-transfecting cells with the heavy chain expression vector pcDNA3.4-17C7H and the light chain expression vector pcDNA3.4-17C7L7 yields the humanized antibody 17C7HL7. Humanized antibody 17C7HL8 was obtained by co-transfecting cells with heavy chain expression vector pcDNA3.4-17C7H4 and light chain expression vector pcDNA3.4-17C7L8 for expression and purification; humanized antibody 17C7H4L was obtained by co-transfecting cells with heavy chain expression vector pcDNA3.4-17C7H12 and light chain expression vector pcDNA3.4-17C7L for expression and purification; humanized antibody 17C7H12L was obtained by co-transfecting cells with heavy chain expression vector pcDNA3.4-17C7H12 and light chain expression vector pcDNA3.4-17C7L for expression and purification. Humanized antibody 17C7H13L was obtained by co-transfecting cells with pcDNA3.4-17C7H13 and light chain expression vector pcDNA3.4-17C7L for expression and purification; similarly, humanized antibody 17C7H17L was obtained by co-transfecting cells with heavy chain expression vector pcDNA3.4-17C7H17 and light chain expression vector pcDNA3.4-17C7L for expression and purification; similarly, humanized antibody 17C7H13L8 was obtained by co-transfecting cells with heavy chain expression vector pcDNA3.4-17C7H13 and light chain expression vector pcDNA3.4-17C7L8 for expression and purification; similarly, humanized antibody 17C7H13L8 was obtained by co-transfecting cells with heavy chain expression vector pcDNA3.4-17C7H13 and light chain expression vector pcDNA3.4-17C7L8 for expression and purification. Humanized antibody 17C7H17L8 was obtained by co-transfecting cells with the heavy chain expression vector pcDNA3.4-17C7H17 and the light chain expression vector pcDNA3.4-17C7L8 for expression and purification. Similarly, humanized antibody 17C7H4L8 was obtained by co-transfecting cells with the heavy chain expression vector pcDNA3.4-17C7H12 and the light chain expression vector pcDNA3.4-17C7L8 for expression and purification.

[0459] Example 10: Affinity detection of different 17C7 humanized antigen-binding protein heavy and light chains with leukocytidine HLGB

[0460] Recombinant leukocytocin HCl was diluted to 1 μg / mL with PBS buffer, and 100 μL was coated into each well of a 96-well plate (Thermo) and incubated overnight at 4°C. The next day, the 96-well plates were removed, washed with PBST (containing 0.5% PBS), and thoroughly dried after each 1-minute wash. 200 μL of PBST containing 5% BSA was added to each sample well, and the plates were blocked at 37°C for 1 hour. The plates were then washed with PBST and the water in the wells was dried. 100 μL of 17C7 was added to each well of the 96-well plate and incubated overnight at 4°C. After washing with PBST, 100 μL of secondary antibody (HRP antibody, Beijing Dingguo Biotechnology Co., Ltd.) was added to each well, and the plates were incubated at 37°C for 1 hour. Wash 5 times with PBST, add 100 μL of Substrate Solution (Invitrogen) to each well, and incubate at 37 °C for 10 min. After stopping the reaction, add 50 μL of 1 mol sulfuric acid to each well and then measure the absorbance at 450 nm using a microplate reader (Multiskcin FC, Thermo).

[0461] The affinity assay results for antigen-binding protein 17C7 and light chain humanized antigen-binding proteins 17C7HL5, 17C7HL6, 17C7HL7, and 17C7HL8 (where the heavy chain variable region is selected from the 17C7 mouse sequence, and the light chain variable regions are selected from the L5, L6, L7, and L8 sequences in Example 8, respectively) with leukocytocin HLB are as follows: Figure 4 As shown, the results indicate that leukocidin HLB has a strong affinity for the light chain humanized monoclonal antibodies 17C7HL5, 17C7HL6, 17C7HL7, and 17C7HL8. Among these, leukocidin HLB binds to EC2 receptors of 17C7HL5. 50 The value was 0.009 μg / mL, indicating that EC bound to 17C7HL6... 50 The value was 0.012 μg / mL, indicating that EC bound to 17C7HL7... 50 The value was 0.016 μg / mL, representing EC binding to 17C7HL8. 50 The value was 0.038 μg / mL.

[0462] The affinity assay results for antigen-binding protein 17C7 and heavy chain humanized antigen-binding proteins 17C7H4L, 17C7H12L, 17C7H13L, and 17C7H17L with leukocidin HLB are as follows: Figure 5 As shown, the results indicate that leukocidin HlgB has a strong affinity for the light chain humanized monoclonal antibodies 17C7H4L, 17C7H12L, 17C7H13L, and 17C7H17L. Among these, leukocidin HlgB binds to EC2 receptors of 17C7H4L. 50The value was 0.014 μg / mL, and the EC bound to 17C7H12L was... 50 The value was 0.03 μg / mL, and the EC bound to 17C7H13L was... 50 The value was 0.02 μg / mL, and the EC bound to 17C7H17L was... 50 The value was 0.17 μg / mL.

[0463] The affinity assay results for antigen-binding protein 17C7 and humanized antigen-binding proteins 17C7H4L8, 17C7H12L8, 17C7H13L8, and 17C7H17L8 with leukocidin HLB are as follows: Figure 6 As shown, the results indicate that leukocidin HlgB has a strong affinity for the light chain humanized monoclonal antibodies 17C7H4L8, 17C7H12L8, 17C7H13L8, and 17C7H17L8. Among these, leukocidin HlgB binds to 17C7H4L8 at EC50. 50 The value was 0.01 μg / mL, and the EC bound to 17C7H12L8 was... 50 The value was 0.019 μg / mL, and the EC bound to 17C7H13L8 was... 50 The value was 0.024 μg / mL, and the EC bound to 17C7H17L8 was [value missing]. 50 The value was 0.035 μg / mL.

[0464] Example 11: Affinity detection of 17C7 humanized antigen-binding protein with various Staphylococcus aureus leukocidins

[0465] Dilute leukocytocin to 1 μg / mL with PBS buffer, and coat each well with 100 μL of the solution in a 96-well plate (Thermo) and incubate overnight at 4°C. The next day, remove the 96-well plate, wash with PBST (containing 0.5% PBS), soaking for 1 min each time and then thoroughly dry. Add 200 μL of PBST containing 5% BSA to each sample well and block at 37°C for 1 h; then wash with PBST and dry the wells. Add 100 μL of 17C7 to each 96-well plate and incubate overnight at 4°C. After removing the 96-well plate, wash with PBST and add 100 μL of secondary antibody (HRP antibody, Beijing Dingguo Biotechnology Co., Ltd.) to each well, and incubate at 37°C for 1 h. Wash 5 times with PBST, add 100 μL of Substrate Solution (Invitrogen) to each well, and incubate at 37 °C for 10 min. After stopping the reaction, add 50 μL of 1 mol sulfuric acid to each well and then measure the absorbance at 450 nm using a microplate reader (Multiskcin FC, Thermo).

[0466] The binding detection results of antigen-binding protein 17C7 and humanized antigen-binding protein 17C7H17L8 to four leukocidins HlgA(A), HlgB(B), LukD(C), and LukF(D) are as follows: Figure 7 As shown, the results indicate that the four antigens mentioned above have a strong affinity for the humanized monoclonal antibody 17C7H17L8. Among them, EC2 binding to HlgA is the most effective. 50 The value was 0.034 μg / mL, and the EC bound to HlgB was... 50 The value was 0.055 μg / mL, and the EC bound to LukD was... 50 The value was 0.036 μg / mL, and the EC binding to LukF was... 50 The value was 0.043 μg / mL.

[0467] Example 12: Replication of a mouse sepsis model induced by Staphylococcus aureus leukocidin HlgAB and evaluation of the pharmacodynamic effects of humanized antigen-binding protein 17C7H17L8.

[0468] C57BL / 6 mice were pre-injected with different doses of antigen-binding protein, and 30 minutes later, 10 μg of Staphylococcus aureus leukocidin HlgAB (containing 5 μg HClA and 5 μg HClB) was injected via the tail vein. Human IgG was used as a control. The survival time of the mice was observed.

[0469] The results of detecting the therapeutic effect of humanized antigen-binding protein 17C7H17L8 using a mouse sepsis model induced by Staphylococcus aureus α-toxin are as follows: Figure 8 As shown. The results indicate that the humanized antigen-binding protein 17C7H17L8 (1.5 mg / kg) described in this application has a complete protective effect.

[0470] Example 13: Replication of a Staphylococcus aureus infection model in mice leading to bacteremia and evaluation of the pharmacodynamic effects of humanized antigen-binding protein 17C7H17L8.

[0471] Staphylococcus aureus USA300 was activated at -80°C, aged two generations in TSA medium, and then inoculated into 2 ml TSB and cultured for 12 h. After washing three times with PBS, 6 × 10⁶ bacteria were injected into the tail vein of mice. 7 CFU. Two hours after infection, the mice were injected with antigen-binding protein, and human IgG was used as a control to observe their survival.

[0472] The results of evaluating the anti-infective pharmacodynamics of humanized antigen-binding protein 17C7H17L8 using a mouse model of Staphylococcus aureus-induced bacteremia are as follows: Figure 9As shown in the figure. The results indicate that the humanized antigen-binding protein 17C7H17L8 has a significant protective effect against Staphylococcus aureus-induced bacteremia in mice. sequence list <110> Xingji Bio (Suzhou) Co., Ltd. <120> Antigen-binding proteins that cross-neutralize Staphylococcus aureus toxins and their uses <130> 0193-PA-008 <160> 66 <170> PatentIn version 3.5 <210> 1 <211> 8 <212> PRT <213> Artificial Sequence <220> <223> HCDR3 <400> 1 Leu Trp Gly Gly Val Leu Asp Tyr 1 5 <210> 2 <211> 17 <212> PRT <213> Artificial Sequence <220> <223> HCDR2 <400> 2 Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe Gln 1 5 10 15 Gly <210> 3 <211> 5 <212> PRT <213> Artificial Sequence <220> <223> HCDR1 <400> 3 Asp Tyr Tyr Met His 1 5 <210> 4 <211> 9 <212> PRT <213> Artificial Sequence <220> <223> LCDR3 <400> 4 Ser Gln Ser Thr His Val Pro Tyr Thr 1 5 <210> 5 <211> 7 <212> PRT <213> Artificial Sequence <220> <223> LCDR2 <400> 5 Arg Val Ser Asn Arg Phe Ser 1 5 <210> 6 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> 17C7L LCDR1 <400> 6 Arg Ser Ser Gln Ser Leu Val Asn Ser Asn Gly Asn Thr Tyr Leu His 1 5 10 15 <210> 7 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4 LCDR1 <400> 7 Arg Ser Ser Gln Ser Leu Val Ala Ser Asn Gly Asn Thr Tyr Leu His 1 5 10 15 <210> 8 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> 17C7L5, 17C7L7 LCDR1 <400> 8 Arg Ser Ser Gln Ser Leu Val Ala Ser Asn Arg Asn Thr Tyr Leu His 1 5 10 15 <210> 9 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> 17C7L6, 17C7L8 LCDR1 <400> 9 Arg Ser Ser Gln Ser Leu Val Ala Ser Asn Arg Ala Thr Tyr Leu His 1 5 10 15 <210> 10 <211> 30 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR1 <400> 10 Glu Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Arg Ser Gly Ala 1 5 10 15 Ser Val Lys Leu Ser Cys Thr Ala Ser Gly Phe Asn Ile Lys 20 25 30 <210> 11 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR2 <400> 11 Trp Val Lys Gln Arg Pro Glu Gln Gly Leu Glu Trp Ile Gly 1 5 10 <210> 12 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR3 <400> 12 Lys Ala Thr Met Thr Ala Asp Thr Ser Ser Asn Thr Ala Tyr Leu Gln 1 5 10 15 Leu Ser Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Phe Cys Asn Ala 20 25 30 <210> 13 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR4 <400> 13 Trp Gly Gln Gly Thr Ser Val Thr Val Ser Ala 1 5 10 <210> 14 <211> 30 <212> PRT <213> Artificial Sequence <220> <223> 17C7H3 H-FR1 <400> 14 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys 20 25 30 <210> 15 <211> 14 <212> PRT <213]] Artificial Sequence <220> <223> 17C7H3, 17C7H4, 17C7H13 H-FR2 <400> 15 Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile Gly 1 5 10 <210> 16 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7H3, 17C7H4, 17C7H12 H-FR3 <400> 16 Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr Met Glu 1 5 10 15 Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys Asn Ala 20 25 30 <210> 17 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 17C7H3, 17C7H4, 17C7H12, 17C7H13, 17C7H17 H-FR4 <400> 17 Trp Gly Gln Gly Thr Leu Val Thr Val Ser Ser 1 5 10 <210> 18 <211> 30 <212> PRT <213> Artificial Sequence <220> <223> 17C7H4, 17C7H12, 17C7H13, 17C7H17 H-FR1 <400> 18 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys 20 25 30 <210> 19 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> 17C7H12, 17C7H17 H-FR2 <400> 19 Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met Gly 1 5 10 <210> 20 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7H13, 17C7H17 H-FR3 <400> 20 Arg Val Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr Met Glu 1 5 10 15 Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys Asn Ala 20 25 30 <210> twenty one <211> twenty three <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR1 <400> twenty one Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly 1 5 10 15 Asp Gln Ala Ser Ile Ser Cys 20 <210> twenty two <211> 15 <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR2 <400> twenty two Trp Tyr Leu Gln Lys Pro Gly Gln Ser Pro Asn Leu Leu Ile Tyr 1 5 10 15 <210> twenty three <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR3 <400> twenty three Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr 1 5 10 15 Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Phe Cys 20 25 30 <210> twenty four <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR4 <400> twenty four Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys Arg 1 5 10 <210> 25 <211> twenty three <212> PRT <213> Artificial Sequence <220> <223> 17C7L4, 17C7L5, 17C7L6 L-FR1 <400> 25 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys 20 <210> 26 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4, 17C7L5, 17C7L6, 17C7L7, 17C7L8 L-FR2 <400> 26 Trp Tyr Gln Gln Arg Pro Gly Gln Pro Pro Arg Leu Leu Ile Tyr 1 5 10 15 <210> 27 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4, 17C7L5, 17C7L6, 17C7L7, 17C7L8 L-FR3 <400> 27 Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr 1 5 10 15 Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys 20 25 30 <210> 28 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4, 17C7L5, 17C7L6, 17C7L7, 17C7L8 L-FR4 <400> 28 Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys Arg 1 5 10 <210> 29 <211> twenty three <212> PRT <213> Artificial Sequence <220> <223> 17C7L7, 17C7L8 L-FR1 <400> 29 Asp Ile Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys 20 <210> 30 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7Hv VH <400> 30 Glu Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Arg Ser Gly Ala 1 5 10 15 Ser Val Lys Leu Ser Cys Thr Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Lys Gln Arg Pro Glu Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Lys Ala Thr Met Thr Ala Asp Thr Ser Ser Asn Thr Ala Tyr 65 70 75 80 Leu Gln Leu Ser Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Phe Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Ser 100 105 110 Val Thr Val Ser Ala 115 <210> 31 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7H3v VH <400> 31 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 32 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7H4v VH <400> 32 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 33 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7H12v VH <400> 33 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 34 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7H13v VH <400> 34 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Val Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 35 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7H17v VH <400> 35 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Val Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser 115 <210> 36 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7Lv VL <400> 36 Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly 1 5 10 15 Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Asn Ser 20 25 30 Asn Gly Asn Thr Tyr Leu His Trp Tyr Leu Gln Lys Pro Gly Gln Ser 35 40 45 Pro Asn Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Phe Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 37 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4v VL <400> 37 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Gly Asn Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 38 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7L5v VL <400> 38 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Asn Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 39 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7L6v VL <400> 39 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Ala Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 40 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7L7v VL <400> 40 Asp Ile Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Asn Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 41 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7L8v VL <400> 41 Asp Ile Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Ala Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 42 <211> 330 <212> PRT <213> Artificial Sequence <220> <223> IgG1 heavy chain constant region <400> 42 Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala Pro Ser Ser Lys 1 5 10 15 Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu Val Lys Asp Tyr 20 25 30 Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly Ala Leu Thr Ser 35 40 45 Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser Gly Leu Tyr Ser 50 55 60 Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu Gly Thr Gln Thr 65 70 75 80 Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr Lys Val Asp Lys 85 90 95 Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr Cys Pro Pro Cys 100 105 110 Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro 115 120 125 Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu Pro Glu Val Thr Cys 130 135 140 Val Val Val Asp Val Ser His Glu Asp Pro Glu Val Lys Phe Asn Trp 145 150 155 160 Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu 165 170 175 Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu 180 185 190 His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn 195 200 205 Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly 210 215 220 Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Arg Glu Glu 225 230 235 240 Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr 245 250 255 Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn 260 265 270 Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp Gly Ser Phe Phe 275 280 285 Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp Gln Gln Gly Asn 290 295 300 Val Phe Ser Cys Ser Val Met His Glu Ala Leu His Asn His Tyr Thr 305 310 315 320 Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 325 330 <210> 43 <211> 106 <212> PRT <213> Artificial Sequence <220> <223> Ig kappa light chain constant region <400> 43 Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu Gln 1 5 10 15 Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe Tyr 20 25 30 Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln Ser 35 40 45 Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser Thr 50 55 60 Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu Lys 65 70 75 80 His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser Pro 85 90 95 Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 100 105 <210> 44 <211> 16 <212> PRT <213> Artificial Sequence <220> <223> 17C7L LCDR1; 17C7L4 LCDR1; 17C7L5, 17C7L7 LCDR1; 17C7L6, 17C7L8 LCDR1 <220> <221> misc_feature <222> (8)..(8) <223> Xaa = Ala or Asn <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Gly or Arg <220> <221> misc_feature <222> (12)..(12) <223> Xaa = Ala or Asn <400> 44 Arg Ser Ser Gln Ser Leu Val Xaa Ser Asn Xaa Xaa Thr Tyr Leu His 1 5 10 15 <210> 45 <211> 30 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR1; 17C7H3 H-FR1; 17C7H4, 17C7H12, 17C7H13, 17C7H17 H-FR1 <220> <221> misc_feature <222> (1)..(1) <223> Xaa = Glu or Gln <220> <221> misc_feature <222> (5)..(5) <223> Xaa = Gln or Val <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Leu or Val <220> <221> misc_feature <222> (12)..(12) <223> Xaa = Lys or Val <220> <221> misc_feature <222> (13)..(13) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (14)..(14) <223> Xaa = Pro or Ser <220> <221> misc_feature <222> (20)..(20) <223> Xaa = Leu or Val <220> <221> misc_feature <222> (23)..(23) <223> Xaa = Lys or Thr <400> 45 Xaa Val Gln Leu Xaa Gln Ser Gly Ala Glu Xaa Xaa Xaa Xaa Gly Ala 1 5 10 15 Ser Val Lys Xaa Ser Cys Xaa Ala Ser Gly Phe Asn Ile Lys 20 25 30 <210> 46 <211> 14 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR2; 17C7H3, 17C7H4, 17C7H13 H-FR2; 17C7H12, 17C7H17 H-FR2 <220> <221> misc_feature <222> (3)..(3) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (5)..(5) <223> Xaa = Ala or Arg <220> <221> misc_feature <222> (7)..(7) <223> Xaa = Glu or Gly <220> <221> misc_feature <222> (13)..(13) <223> Xaa = Ile or Met <400> 46 Trp Val Xaa Gln Xaa Pro Xaa Gln Gly Leu Glu Trp Xaa Gly 1 5 10 <210> 47 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR3; 17C7H3, 17C7H4, 17C7H12 H-FR3; 17C7H13, 17C7H17 H-FR3 <220> <221> misc_feature <222> (1)..(1) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (2)..(2) <223> Xaa = Ala or Val <220> <221> misc_feature <222> (10)..(10) <223> Xaa = Ile or Ser <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Asn or Ser <220> <221> misc_feature <222> (15)..(15) <223> Xaa = Leu or Met <220> <221> misc_feature <222> (16)..(16) <223> Xaa = Glu or Gln <220> <221> misc_feature <222> (19)..(19) <223> Xaa = Arg or Ser <220> <221> misc_feature <222> (21)..(21) <223> Xaa = Arg or Thr <220> <221> misc_feature <222> (23)..(23) <223> Xaa = Asp or Glu <220> <221> misc_feature <222> (29)..(29) <223> Xaa = Phe or Tyr <400> 47 Xaa Xaa Thr Met Thr Ala Asp Thr Ser Xaa Xaa Thr Ala Tyr Xaa Xaa 1 5 10 15 Leu Ser Xaa Leu Xaa Ser Xaa Asp Thr Ala Val Tyr Xaa Cys Asn Ala 20 25 30 <210> 48 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 17C7H H-FR4; 17C7H3, 17C7H4, 17C7H12, 17C7H13, 17C7H17 H-FR4 <220> <221> misc_feature <222> (6)..(6) <223> Xaa = Leu or Ser <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Ala or Ser <400> 48 Trp Gly Gln Gly Thr Xaa Val Thr Val Ser Xaa 1 5 10 <210> 49 <211> twenty three <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR1; 17C7L4, 17C7L5, 17C7L6 L-FR1; 17C7L7, 17C7L8 L-FR1 <220> <221> misc_feature <222> (2)..(2) <223> Xaa = Ile or Val <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Leu or Ser <220> <221> misc_feature <222> (14)..(14) <223> Xaa = Ser or Thr <220> <221> misc_feature <222> (17)..(17) <223> Xaa = Asp or Gln <220> <221> misc_feature <222> (18)..(18) <223> Xaa = Pro or Gln <400> 49 Asp Xaa Val Met Thr Gln Thr Pro Leu Ser Xaa Pro Val Xaa Leu Gly 1 5 10 15 Xaa Xaa Ala Ser Ile Ser Cys 20 <210> 50 <211> 15 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4, 17C7L5, 17C7L6, 17C7L7, 17C7L8 L-FR2; 17C7L L-FR2 <220> <221> misc_feature <222> (3)..(3) <223> Xaa = Leu or Gln <220> <221> misc_feature <222> (5)..(5) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (9)..(9) <223> Xaa = Pro or Ser <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Asn or Arg <400> 50 Trp Tyr Xaa Gln Xaa Pro Gly Gln Xaa Pro Xaa Leu Leu Ile Tyr 1 5 10 15 <210> 51 <211> 32 <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR3; 17C7L4, 17C7L5, 17C7L6, 17C7L7, 17C7L8 L-FR3 <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Ala or Ser <220> <221> misc_feature <222> (27)..(27) <223> Xaa = Leu or Val <220> <221> misc_feature <222> (31)..(31) <223> Xaa = Phe or Tyr <400> 51 Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Xaa Gly Thr Asp Phe Thr 1 5 10 15 Leu Lys Ile Ser Arg Val Glu Ala Glu Asp Xaa Gly Val Tyr Xaa Cys 20 25 30 <210> 52 <211> 11 <212> PRT <213> Artificial Sequence <220> <223> 17C7L L-FR4; 17C7L4, 17C7L5, 17C7L6, 17C7L7, 17C7L8 L-FR4 <220> <221> misc_feature <222> (3)..(3) <223> Xaa = Gly or Gln <400> 52 Phe Gly Xaa Gly Thr Lys Leu Glu Ile Lys Arg 1 5 10 <210> 53 <211> 117 <212> PRT <213> Artificial Sequence <220> <223> 17C7H VH; 17C7H3 VH; 17C7H4 VH; 17C7H12 VH; 17C7H13 VH; 17C7H17 VH <220> <221> misc_feature <222> (1)..(1) <223> Xaa = Glu or Gln <220> <221> misc_feature <222> (5)..(5) <223> Xaa = Gln or Val <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Leu or Val <220> <221> misc_feature <222> (12)..(12) <223> Xaa = Lys or Val <220> <221> misc_feature <222> (13)..(13) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (14)..(14) <223> Xaa = Pro or Ser <220> <221> misc_feature <222> (20)..(20) <223> Xaa = Leu or Val <220> <221> misc_feature <222> (23)..(23) <223> Xaa = Lys or Thr <220> <221> misc_feature <222> (38)..(38) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (40)..(40) <223> Xaa = Ala or Arg <220> <221> misc_feature <222> (42)..(42) <223> Xaa = Glu or Gly <220> <221> misc_feature <222> (48)..(48) <223> Xaa = Ile or Met <220> <221> misc_feature <222> (67)..(67) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (68)..(68) <223> Xaa = Ala or Val <220> <221> misc_feature <222> (76)..(76) <223> Xaa = Ile or Ser <220> <221> misc_feature <222> (77)..(77) <223> Xaa = Asn or Ser <220> <221> misc_feature <222> (81)..(81) <223> Xaa = Leu or Met <220> <221> misc_feature <222> (82)..(82) <223> Xaa = Glu or Gln <220> <221> misc_feature <222> (85)..(85) <223> Xaa = Arg or Ser <220> <221> misc_feature <222> (87)..(87) <223> Xaa = Arg or Thr <220> <221> misc_feature <222> (89)..(89) <223> Xaa = Asp or Glu <220> <221> misc_feature <222> (95)..(95) <223> Xaa = Phe or Tyr <220> <221> misc_feature <222> (112)..(112) <223> Xaa = Leu or Ser <220> <221> misc_feature <222> (117)..(117) <223> Xaa = Ala or Ser <400> 53 Xaa Val Gln Leu Xaa Gln Ser Gly Ala Glu Xaa Xaa Xaa Xaa Gly Ala 1 5 10 15 Ser Val Lys Xaa Ser Cys Xaa Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Xaa Gln Xaa Pro Xaa Gln Gly Leu Glu Trp Xaa 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Xaa Xaa Thr Met Thr Ala Asp Thr Ser Xaa Xaa Thr Ala Tyr 65 70 75 80 Xaa Xaa Leu Ser Xaa Leu Xaa Ser Xaa Asp Thr Ala Val Tyr Xaa Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Xaa 100 105 110 Val Thr Val Ser Xaa 115 <210> 54 <211> 113 <212> PRT <213> Artificial Sequence <220> <223> 17C7L VL; 17C7L4 VL; 17C7L5 VL; 17C7L6 VL; 17C7L7 VL; 17C7L8 VL <220> <221> misc_feature <222> (2)..(2) <223> Xaa = Ile or Val <220> <221> misc_feature <222> (11)..(11) <223> Xaa = Leu or Ser <220> <221> misc_feature <222> (14)..(14) <223> Xaa = Ser or Thr <220> <221> misc_feature <222> (17)..(17) <223> Xaa = Asp or Gln <220> <221> misc_feature <222> (18)..(18) <223> Xaa = Pro or Gln <220> <221> misc_feature <222> (31)..(31) <223> Xaa = Ala or Asn <220> <221> misc_feature <222> (34)..(34) <223> Xaa = Gly or Arg <220> <221> misc_feature <222> (35)..(35) <223> Xaa = Ala or Asn <220> <221> misc_feature <222> (42)..(42) <223> Xaa = Leu or Gln <220> <221> misc_feature <222> (44)..(44) <223> Xaa = Lys or Arg <220> <221> misc_feature <222> (48)..(48) <223> Xaa = Pro or Ser <220> <221> misc_feature <222> (50)..(50) <223> Xaa = Asn or Arg <220> <221> misc_feature <222> (72)..(72) <223> Xaa = Ala or Ser <220> <221> misc_feature <222> (88)..(88) <223> Xaa = Leu or Val <220> <221> misc_feature <222> (92)..(92) <223> Xaa = Phe or Tyr <220> <221> misc_feature <222> (105)..(105) <223> Xaa = Gly or Gln <400> 54 Asp Xaa Val Met Thr Gln Thr Pro Leu Ser Xaa Pro Val Xaa Leu Gly 1 5 10 15 Xaa Xaa Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Xaa Ser 20 25 30 Asn Xaa Xaa Thr Tyr Leu His Trp Tyr Xaa Gln Xaa Pro Gly Gln Xaa 35 40 45 Pro Xaa Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Xaa Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Xaa Gly Val Tyr Xaa Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Xaa Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg <210> 55 <211> 447 <212> PRT <213> Artificial Sequence <220> <223> 17C7 chimeric antibody heavy chain (17C7H) <400> 55 Glu Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Arg Ser Gly Ala 1 5 10 15 Ser Val Lys Leu Ser Cys Thr Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Lys Gln Arg Pro Glu Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Lys Ala Thr Met Thr Ala Asp Thr Ser Ser Asn Thr Ala Tyr 65 70 75 80 Leu Gln Leu Ser Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Phe Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Ser 100 105 110 Val Thr Val Ser Ala Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 115 120 125 Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys 130 135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn 195 200 205 Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His 210 215 220 Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val 225 230 235 240 Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu 245 250 255 Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 260 265 270 Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 275 280 285 Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser 290 295 300 Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys 305 310 315 320 Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile 325 330 335 Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro 340 345 350 Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu 355 360 365 Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 370 375 380 Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser 385 390 395 400 Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg 405 410 415 Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu 420 425 430 His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly Lys 435 440 445 <210> 56 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> 17C7 chimeric antibody light chain (17C7L) <400> 56 Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly 1 5 10 15 Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Asn Ser 20 25 30 Asn Gly Asn Thr Tyr Leu His Trp Tyr Leu Gln Lys Pro Gly Gln Ser 35 40 45 Pro Asn Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Phe Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe 130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 210 215 <210> 57 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> 17C7H3 heavy chain <400> 57 Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 115 120 125 Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys 130 135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn 195 200 205 Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His 210 215 220 Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val 225 230 235 240 Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu 245 250 255 Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 260 265 270 Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 275 280 285 Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser 290 295 300 Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys 305 310 315 320 Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile 325 330 335 Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro 340 345 350 Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu 355 360 365 Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 370 375 380 Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser 385 390 395 400 Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg 405 410 415 Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu 420 425 430 His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 <210> 58 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> 17C7H4 heavy chain <400> 58 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 115 120 125 Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys 130 135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn 195 200 205 Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His 210 215 220 Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val 225 230 235 240 Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu 245 250 255 Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 260 265 270 Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 275 280 285 Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser 290 295 300 Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys 305 310 315 320 Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile 325 330 335 Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro 340 345 350 Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu 355 360 365 Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 370 375 380 Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser 385 390 395 400 Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg 405 410 415 Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu 420 425 430 His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 <210> 59 <211> 445 <212> PRT <213> Artificial Sequence <220> <223> 17C7H12 heavy chain <400> 59 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Ala Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu Ala 115 120 125 Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys Leu 130 135 140 Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser Gly 145 150 155 160 Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser Ser 165 170 175 Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser Leu 180 185 190 Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn Thr 195 200 205 Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His Thr 210 215 220 Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val Phe 225 230 235 240 Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu Pro 245 250 255 Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu Val 260 265 270 Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys Thr 275 280 285 Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser Val 290 295 300 Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys Cys 305 310 315 320 Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile Ser 325 330 335 Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro Pro 340 345 350 Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu Val 355 360 365 Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn Gly 370 375 380 Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser Asp 385 390 395 400 Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg Trp 405 410 415 Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu His 420 425 430 Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 <210> 60 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> 17C7H13 heavy chain <400> 60 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Ile 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Val Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 115 120 125 Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys 130 135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn 195 200 205 Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His 210 215 220 Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val 225 230 235 240 Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu 245 250 255 Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 260 265 270 Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 275 280 285 Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser 290 295 300 Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys 305 310 315 320 Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile 325 330 335 Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro 340 345 350 Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu 355 360 365 Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 370 375 380 Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser 385 390 395 400 Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg 405 410 415 Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu 420 425 430 His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 <210> 61 <211> 446 <212> PRT <213> Artificial Sequence <220> <223> 17C7H17 heavy chain <400> 61 Glu Val Gln Leu Val Gln Ser Gly Ala Glu Val Lys Lys Pro Gly Ala 1 5 10 15 Ser Val Lys Val Ser Cys Lys Ala Ser Gly Phe Asn Ile Lys Asp Tyr 20 25 30 Tyr Met His Trp Val Arg Gln Ala Pro Gly Gln Gly Leu Glu Trp Met 35 40 45 Gly Trp Ile Asp Pro Glu Asn Asp Ile Gly Glu Tyr Ala Pro Lys Phe 50 55 60 Gln Gly Arg Val Thr Met Thr Ala Asp Thr Ser Ile Ser Thr Ala Tyr 65 70 75 80 Met Glu Leu Ser Arg Leu Arg Ser Asp Asp Thr Ala Val Tyr Tyr Cys 85 90 95 Asn Ala Leu Trp Gly Gly Val Leu Asp Tyr Trp Gly Gln Gly Thr Leu 100 105 110 Val Thr Val Ser Ser Ala Ser Thr Lys Gly Pro Ser Val Phe Pro Leu 115 120 125 Ala Pro Ser Ser Lys Ser Thr Ser Gly Gly Thr Ala Ala Leu Gly Cys 130 135 140 Leu Val Lys Asp Tyr Phe Pro Glu Pro Val Thr Val Ser Trp Asn Ser 145 150 155 160 Gly Ala Leu Thr Ser Gly Val His Thr Phe Pro Ala Val Leu Gln Ser 165 170 175 Ser Gly Leu Tyr Ser Leu Ser Ser Val Val Thr Val Pro Ser Ser Ser 180 185 190 Leu Gly Thr Gln Thr Tyr Ile Cys Asn Val Asn His Lys Pro Ser Asn 195 200 205 Thr Lys Val Asp Lys Arg Val Glu Pro Lys Ser Cys Asp Lys Thr His 210 215 220 Thr Cys Pro Pro Cys Pro Ala Pro Glu Leu Leu Gly Gly Pro Ser Val 225 230 235 240 Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Tyr Ile Thr Arg Glu 245 250 255 Pro Glu Val Thr Cys Val Val Val Asp Val Ser His Glu Asp Pro Glu 260 265 270 Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu Val His Asn Ala Lys 275 280 285 Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg Val Val Ser 290 295 300 Val Leu Thr Val Leu His Gln Asp Trp Leu Asn Gly Lys Glu Tyr Lys 305 310 315 320 Cys Lys Val Ser Asn Lys Ala Leu Pro Ala Pro Ile Glu Lys Thr Ile 325 330 335 Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr Thr Leu Pro 340 345 350 Pro Ser Arg Glu Glu Met Thr Lys Asn Gln Val Ser Leu Thr Cys Leu 355 360 365 Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ser Asn 370 375 380 Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val Leu Asp Ser 385 390 395 400 Asp Gly Ser Phe Phe Leu Tyr Ser Lys Leu Thr Val Asp Lys Ser Arg 405 410 415 Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His Glu Ala Leu 420 425 430 His Asn His Tyr Thr Gln Lys Ser Leu Ser Leu Ser Pro Gly 435 440 445 <210> 62 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> 17C7L4 light chain <400> 62 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Gly Asn Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe 130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 210 215 <210> 63 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> 17C7L5 light chain <400> 63 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Asn Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe 130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 210 215 <210> 64 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> 17C7L6 light chain <400> 64 Asp Val Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Ala Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe 130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 210 215 <210> 65 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> 17C7L7 light chain <400> 65 Asp Ile Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Asn Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe 130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 210 215 <210> 66 <211> 219 <212> PRT <213> Artificial Sequence <220> <223> 17C7L8 light chain <400> 66 Asp Ile Val Met Thr Gln Thr Pro Leu Ser Ser Pro Val Thr Leu Gly 1 5 10 15 Gln Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Leu Val Ala Ser 20 25 30 Asn Arg Ala Thr Tyr Leu His Trp Tyr Gln Gln Arg Pro Gly Gln Pro 35 40 45 Pro Arg Leu Leu Ile Tyr Arg Val Ser Asn Arg Phe Ser Gly Val Pro 50 55 60 Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Lys Ile 65 70 75 80 Ser Arg Val Glu Ala Glu Asp Val Gly Val Tyr Tyr Cys Ser Gln Ser 85 90 95 Thr His Val Pro Tyr Thr Phe Gly Gln Gly Thr Lys Leu Glu Ile Lys 100 105 110 Arg Thr Val Ala Ala Pro Ser Val Phe Ile Phe Pro Pro Ser Asp Glu 115 120 125 Gln Leu Lys Ser Gly Thr Ala Ser Val Val Cys Leu Leu Asn Asn Phe 130 135 140 Tyr Pro Arg Glu Ala Lys Val Gln Trp Lys Val Asp Asn Ala Leu Gln 145 150 155 160 Ser Gly Asn Ser Gln Glu Ser Val Thr Glu Gln Asp Ser Lys Asp Ser 165 170 175 Thr Tyr Ser Leu Ser Ser Thr Leu Thr Leu Ser Lys Ala Asp Tyr Glu 180 185 190 Lys His Lys Val Tyr Ala Cys Glu Val Thr His Gln Gly Leu Ser Ser 195 200 205 Pro Val Thr Lys Ser Phe Asn Arg Gly Glu Cys 210 215

Claims

1. An isolated antigen-binding protein that specifically binds to Staphylococcus aureus leukocidin, wherein the antibody heavy chain variable region VH of the antigen-binding protein comprises HCDR1, HCDR2, and HCDR3, and the antibody light chain variable region VL of the antigen-binding protein comprises LCDR1, LCDR2, and LCDR3, wherein HCDR1 is shown in SEQ ID NO:3, HCDR2 is shown in SEQ ID NO:2, and HCDR3 is shown in SEQ ID NO:

1. The LCDR1 is shown in SEQ ID NO:6, the LCDR2 is shown in SEQ ID NO:5, and the LCDR3 is shown in SEQ ID NO:4, or The LCDR1 is shown in SEQ ID NO:8, the LCDR2 is shown in SEQ ID NO:5, and the LCDR3 is shown in SEQ ID NO:4, or The LCDR1 is shown as SEQ ID NO:9, the LCDR2 is shown as SEQ ID NO:5, and the LCDR3 is shown as SEQ ID NO:

4.

2. The antigen-binding protein that specifically binds to Staphylococcus aureus leukocidin as described in claim 1, characterized in that, The VH is as shown in SEQ ID NO: 30 and the VL is as shown in SEQ ID NO: 36, or The VH is as shown in SEQ ID NO: 30 and the VL is as shown in SEQ ID NO: 38, or The VH is as shown in SEQ ID NO: 30 and the VL is as shown in SEQ ID NO: 39, or The VH is as shown in SEQ ID NO: 30 and the VL is as shown in SEQ ID NO: 40, or The VH is as shown in SEQ ID NO: 30 and the VL is as shown in SEQ ID NO: 41, or The VH is as shown in SEQ ID NO: 32 and the VL is as shown in SEQ ID NO: 36, or The VH is as shown in SEQ ID NO: 33 and the VL is as shown in SEQ ID NO: 36, or The VH is as shown in SEQ ID NO: 34 and the VL is as shown in SEQ ID NO: 36, or The VH is as shown in SEQ ID NO: 35 and the VL is as shown in SEQ ID NO: 36, or The VH is as shown in SEQ ID NO: 32 and the VL is as shown in SEQ ID NO: 41, or The VH is as shown in SEQ ID NO: 33 and the VL is as shown in SEQ ID NO: 41, or The VH is as shown in SEQ ID NO: 34 and the VL is as shown in SEQ ID NO: 41, or The VH is shown as SEQ ID NO: 35 and the VL is shown as SEQ ID NO:

41.

3. A nucleic acid molecule encoding an antigen-binding protein that specifically binds to Staphylococcus aureus leukocidin isolated according to any one of claims 1 or 2.

4. A vector comprising the nucleic acid molecule of claim 3.

5. A cell comprising the nucleic acid molecule of claim 3 or the vector of claim 4.

6. A pharmaceutical composition comprising the isolated antigen-binding protein that specifically binds to Staphylococcus aureus leukocidin as described in claim 1 or 2, the nucleic acid molecule as described in claim 3, the carrier as described in claim 4, and / or the cell as described in claim 5, and optionally a pharmaceutically acceptable carrier.

7. Use in the preparation of a medicament of the isolated antigen-binding protein specifically binding to Staphylococcus aureus leukocidin as described in claim 1 or 2, the nucleic acid molecule as described in claim 3, the carrier as described in claim 4, the cell as described in claim 5, and / or the pharmaceutical composition as described in claim 6, for the prevention and / or treatment of diseases and / or symptoms caused by Staphylococcus aureus leukocidin.

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